Study on Liquid Culture of Poria Cocos Triterpenes and Its Fermented Production
|School||Anhui Agricultural University|
|Course||Nutrition and Food Hygiene|
|Keywords||Poria cocos Liquid Fermentation Triterpenes RSM Volatile components|
Poria cocos is called as dodder, fuling, loose potatoes and paneling, which is the aridsclerotia of Poria cocos（Sch.Wolf） and a kind of fungus belonging to polyporaceae. It isadmitted in Chinese Pharmacopeia （2005）. In recent years, foreign and Chinese scholarshave found that the triterpenes of Poria cocos have significant pharmacological functions,such as anticancer activities, anti-bacteria, anti-inflammatory, anti-tumor, enhanceimmunity, anti-convulsant and so on. In the study, the culture conditions of Poria cocoswere optimized to increase the total triterpenes production. Meanwhile, the fermentationproducts of Poria cocos cultured were extracted and analyzed.The spectrophotometric quantitative method was established to determine the contentof total triterpenes of Poria cocos in submerged culture by using vanillin-perchloric acid asthe developer. The reaction conditions were optimized. Taking ursolic acid as the standard,5%vanillin-acetic acid （0.2mL） and perchloric acid （0.8mL） as the developer, after beingincubated at60°C for20min the Abs value of sample solution was determined at548nm.This method operated conveniently, and the result was stable and repeatable.In order to increase the total triterpenes production, the culture conditions of Poriacocos were optimized by the single factor experiment. The optimized medium was showedas following: glucose30.0g L-1, peptone10.0g L-1, KH2PO42.0g L-1, MgSO41.0g L-1,corn meal15.0g L-1, VB10.006g L-1. The optimized result was temperature of26.0,rotation speed of130r min-1, initial pH of5.5, inoculum size of6.0%, broth content of50mL. On the basis of a single factor test, the initial pH, VB1, rotation speed andtemperature as the main factor were screened through the Plackett-Burman design,according to the Plackett-Burman experimental results and the coefficient estimate of eachfactor, they determine the effect of changes in step size and the best approximation of thevalue of the fastest growing region, which has been the center of the response surfaceanalysis. RSM was used to obtain the optimal results: initial pH5.45,VB17.3mg L-1,rotation speed132r min-1, culture temperature26.6. The result showed that the triterpeneproduction under the optimal culture condition reached to7.06percent. It was increased to35percent as compared with the pre-optimized yield.The extraction technology of total triterpenes was researched from Poria cocosmycelia by ultrasonic-assist. The extraction condition of total triterpenes in the mycelia ofPoria cocos was optimized by the method of the single factor experiment and Box-Behnken design test. The results indicated that the optimal extraction conditions wereas follows: ultrasonic power290W, ultrasonic extraction time61min, and ultrasonicextraction temperature41, liquid-solid ratio1/50g mL-1. With the optimized technology,the content of total triterpenes in the final product is7.59percent.Under LC-MS condition, pachymic acid was tentatively authenticated in the myceliaof Poria cocos based on the accurate molecular weight by mass spectrometry.49compounds were identified in the mycelia of Poria cocos, the main constituents inessential oil of Poria cocos mycelia were alcohols（6）, ketones（5）, nitrogens（6）, olefins（4）,acids（7）, aromatics（3）, alkane hydrocarbons（14）, ester（1）, terpene hydrocarbons（1）,terpenoids（1） and other（1）.49compounds were identified in the fermentation liquid, themain constituents in essential oil of fermentation liquid were alcohols（4）, esters（8）,ketones（8）, nitrogens（5）, aromatics（2）, furan（1）, olefin（1）, alkyne（1）, ether （1）and alkanehydrocarbons（18）.