Isolation And Identification Of Secondary Metabolites Produced By Bacillus Pumilus

Rice blast is one kind of the major diseases which is widely distributed all over the word. The disease which restrict the development of the rice production mostly is rice blast. Rice blast can cause rice yields to fall roughly 15-30%. According to some statistics, the reduction of rice caused by rice blast can feed 60 million people each year. Food production caused by the rice blast has become an important economic issue.The main measure against the rice blast is using pesticide and growing disease-resistant varieties. But it is hard to against rice blast by growing disease-resistant varieties because the rice blast genetic background is complex. Although chemical control have many advantages like low cost and effective, overuse of pesticides not only pollute the environment but also make rice blast become more drug resistance. Biological control is a bioeffector-method of controlling plant diseases using other living organisms.We isolated a bacterial strain Bp6221 which has strong antagonistic activity against rice blast from the fruit of Osmanthus.Bp6221 was identified as bacillus pumillus by morphological features and molecular genetic analyses. Extraction experiments showed that the active compound which has strong polarity can soluble in polar solvents such as methanol and water. The culture filtrate of Bp6221 has obvious inhibitory effect on rice blast. The half concentration of culture filtrate is 0.0025mL/mL. The temperature experiments show that the active compound has good thermal stability. It can remain the same activity even treated at 100? for 30 min. It also has good acid-resistance that can remain the same activity after treated in heavily acidic conditions for 24 h.We isolated and purified the active compound using the method of precipitation and chromatography. Firstly, we precipitated protein with acetone. Secondly, we isolated and purified the active compound by running a column. Finally, we collected the active compound which has a good purity by using HPLC. The result showed that there have four kind active compounds. Their retention time is 14.112,14.188,19.140 and 19.384 min. After isolation and purification, the four compounds were identified by LC-MASS. The molecular weight of the four compounds were 150.68,149.78,177.68 and 292.79.As one kind of Bacillus pumilus, Bp6221 has good ability to adapt the environment and been widely found soil, water, air, plants and animals. As a new kind of biological pesticide, the secondary metabolite of Bp6221 has a promising prospect because of its excellent activity, small molecular weight, thermal and acid stability.