Optimization Of Liquid Fermentation Process And Separation And Purification Of Pigment From Bamboo And Yellow

Shiraia bambusicola is an important medicinal fungi with the value of development and application,and it contains a variety of active natural ingredients. Many of the active ingredients make Shiraia bambusicola have a cough expectorant, stasis dysmenorrhea, blood and pain, restore tissue function, enhance physical fitness and other effects. With the deepening of research to its mechanism of pharmacological effects, the medicinal value of Shiraia bambusicola is increasingly prominent, its development and application prospects are very attractive. But because of the use of the fruiting body of Shiraia is limited by its resources,it can not meet the large-scale development and utilization, and its secondary metabolites may be related to its endophytes. In this paper, while Shiraia bambusicola isolate—Fusarium chlamydosporum act as the tester strain,we studied its liquid fermentation conditions, extraction and separation of red pigment in Fusarium chlamydosporum, pigment and pigment stability test and portion of functional tests about the crude extracts. Details are as follows:(1) The optimal fermentation condition and mediun are selected by mono-factor design and orthogonal experiment. Medium components contains carbon source and its concentration, nitrogen sources, metal ions and their concentration, pH, While optimal fermentation condition conditions incubation temperature, incubation time, medium volume and shaking speed.Throughout this progress,we make biomass and pigment production as the indicator.The final liquid fermentation medium and optimal culture conditions are as follows:20%of potato, glucose3%, MgSO4·7H2O0.03%, pH4,temperature28℃, shaking speed180rpm, liquid volume120mL/250mL, incubation time6d. After this final verification of fermentation conditions, the absorbance of the dye solution resch to0.853,which is2.35times to the before optimization level0.363;the content of crude pigment improved from2g/L to4.5g/L;and the biomass improves from9g/L to16.3g/L,increased1.8times.(2) After investigated of extraction solvent, extraction time and solid-liquid ratio of the red pigment,The results are as follows:extraction solvent, extraction time,solid-liquid ratio, respectively are:dichloromethane,2h,1:30.In addition,compared with standing leaching,the ultrasonic extraction not only have high extraction efficiency,it can also increase the amount of red pigment in Fusarium chlamydosporum.Ultrasonic extraction2h can achieve the effect of standing extraction20min.It can save significant amount of time.After the red pigment who was produced by test Fusarium chlamydosporum was isolated through chromuatography on silica gel column,we obtain the pigment sample with a certain purity, by leucomethylene blue color reaction, We can deny that it is anthraquinone dyes,and it is more possibly to be benzoquinone or naphthoquinone class.And UV absorption peak of258nm is also a characteristic absorption peak of naphthoquinone.After MS detection of the pigment samples, we obtained a molecular weight of594.(3) After stability studies of Fusarium chlamydosporum,we found that the red pigment is very unstable to strong light, but temperature has little effect on its stability.Through the antibacterial test of the red pigment,we found that, the red pigment of1mg/mL、0.5mg/mL、0.25mg/mL has a strong inhibitory effect on Bacillus subtilis and E.coli, have little inhibitory effect on Staphylococcus aureus and yeasts,but have no effect on the Green mold.Inhibition of tyrosinase activity test results show that the pigment has good inhibitory activity of tyrosinase,while the concentration of the pigment and the positive control arbutin are all lmg/mL,their tyrosinase activity inhibition rate are respectively64%and79%.