Biosynthesis Of Anisic Acid By Trans-Anethole And Its Techniques Optimazation

The screening of strains degradating and transformating trans-anethole into anisic acid was investigated, as well as the fermentation optimization of the strain, and the key intermediate in degradation. Meanwhile, in view of trans-anethole biodegradation and transformation system, the quantitative analysis method for it by ultraviolet spectrophotometry and high performance liquid chromatography was established. And the main achievements are as follows:(1)A method for analysising by ultraviolet spectrophtometry the system of trans-anethole and anisic acid was established. Under certain conditions, the absorption values at250nm and268.50of trans-anethole, anisic acid and their mixture were proportional with their total molar concentration and the mole percentage of trans-anethole, respectively. With the simulation technology, a rapid quantitative method analysising for the system was established.(2) A method was set up to determinate simultaneously the concentrations of trans-anethole, anise alcohol, anisaldehyde and ansic acid. Moreover the optimum chromatographic conditions were260nm as UV wavelength, acetonitrile-water (70:30) as mobile phase,1mL-min-1as flow rate, and25℃as column temperature. And the experiment results showed that the accurate and reliable method could be used to monitor and detecte the trans-anethole biotransformation products.(3) A strain biodegradating and transformating trans-anethole was screened. First of all,189strains were screened from the anisic trees branches, leaves, fruit and soil in Guangxi University farm and GaoFeng forest farm. Finally, a target strain ZZ-1which could biosynthesis anisic acid at molar ratio9.21%was obtained and identified as mould.(4)The medium composition, fermentation and transformation condition of strain ZZ-1were optimizated. The optimum medium formula was as follows: glucose25g·L-1, yeast extract2.5g·L-1, ammonium chloride5.0g·9L-1, NaCl0.3g·L-1, MgSO4·7H2O0.3g·L-1, K2HPO4·3H2O1.2g-L-1,FeSO4·7H2O0.01g·L-1; and the optimum culture and transformation conditions were initial pH6, temperature30℃, culture and transform time72hours. After optimization, the molar conversion rate of anisic acid increased from9.21%to19.10%.(5)The key intermediate in trans-anethole biobiological degradation and transformation system was investigated. With chromatography separation, GC-MS and IR detection, some intermediate such as1-(4-methoxyphenyl)-1,2-epoxy propane,1-(4-Methoxyphenyl)-1,2-propanediol and anisic alcohol were found, which could provide basis reference for metabolic pathway analysis of trans-anethole biosynthesising anisic acid and its intermediate’s research and application.