Novel Biotransformation Of Podophyllotoxin To Podophyllic Acid And Picropodophyllotoxin

For the first time, a biotransformation process of podophyllotoxin to produce podophyllic acid and picropodophyllotoxin was developed in this work. High substrate conversion and product concentration were obtained through process optimization.First, the novel biotransformation process of podophyllotoxin to produce podophyllic acid and picropodophyllotoxin was developed in this work. Eight bacteria which could modify the structure of podophyllotoxin were screened out from the tested fourteen bacteria, and they were Pseudomonas aeruginosa CCTCC AB 93066, Bacillus sp. CCTCC AB95018, Rhodococcus erythropolis CGMCC 1.2362, Bacillus fusiformis CICC 20463, Corynebacterium pekinenese CCTCC AB 97005, Bacillus subtilis CCTCC AB93174, Erwinia uredovora CGMCC1.1215 and Pseudomonas geniculata CCTCC AB93074. The highest conversion of podophyllotoxin (i.e., 70.2%±8.0%) was obtained when P. aeruginosa was used as biocatalyst, so P. aeruginosa was selected as a typical biocatalyst in the following study. Product (1) and (2) were separated through D312 macroporous resin and sephadex LH-20 gel column chromatograph. On the basis of 1H NMR,13C NMR, ESI-MS and Elemental Analysis, product (1) and (2) were identified as podophyllic acid (1) and picropodophyllotoxin (2), respectively. For the first time, podophyllotoxin was biotransformed into its hydrolytic derivate (i.e., podophyllic acid), and podophyllic acid showed impressive depressant effect on ovarian cancer and melanoma.Then, this work optimized the novel biotransformation process of podophyllotoxin to produce podophyllic acid by P. aeruginosa. Firstly, the biotransformation process was significantly affected by medium composition. 5 g/L of yeast extract and 5 g/L of peptone were favorable for podophyllic acid production (i.e., 25.3±3.7 mg/L), while not beneficial for the cell growth of P. aeruginosa. This indicated that the accumulation of podophyllic acid was not corresponded well to the cell growth of P. aeruginosa. 0 g/L of sucrose was beneficial for podophyllic acid production (i.e., 34.3±3.9 mg/L), which led to high podophyllotoxin conversion (i.e., 98.2%±0.1%). 1 g/L of NaCl was the best for podophyllic acid production (i.e., 47.6±4.0 mg/L). Secondly, the production of podophyllic acid was significantly enhanced by fed-batch biotransformation. When 100 mg/L of substrate was added to the biotransformation system at the 4, 10 and 25 hours, respectively, podophyllic acid concentration reached 99.9±12.3 mg/L, enhanced by 284% comparing to one-time addition (i.e., 26.0±2.1 mg/L).Podophyllotoxin can be used as leading compound to generate new compounds with diversiform and novel structure by structure modification and alteration. Pseudomonas aeruginosa and other seven strains of bacteria were successfully applied to biotransform podophyllotoxin in the paper, yielding podophyllic acid and picropodophyllotoxin, which will provide guidance for the penetrating study on biotransformation of podophyllotoxin, and give reference to the biotransformation study of other bioactive natural leading compounds.