The Transformation Of Trans-Cinnamic Acid Into L-phenylalanine By Phenylalanine Ammonia-lyase

L-phenylalanine(L-Phe), an essential aromatic amino acid, has been widely used in food processing, feed additive and pharmaceutical industry. Aspartame, an important sweetener, produced from L-phenylalanine was permitted to sell by many countries, which greatly improved the demand of L-phenylalanine.An enzymatic method using phenylalanine ammonia-lyase(PAL) of microbe for the transformation of trans-cinnamic acid(t-Ca) to L-phenylalanine was investigated in the paper, majored in incubation of strains, conditions of producing L-phenylalanine ammonia-lyase and conditions of transformation.The yeast strain with high activity of producing phenylalanine ammonia-lyase, Rhodotorula glutinis JM432, was screened from soil. ZW112, with high yield of phenylalanine ammonia-lyase, was a mutant strain of JM432 treated by UV. It is proved that its higher phenylalanine ammonia-lyase producing activity could be inherited after several subcultures.The optimum conditions for seed culture were as follows:10g/L corn steep liquor,initial pH 5.5 and 50mL medium in 250mL fermentation flask incubated at 32℃,140r/min for 34 hours.The optimal medium for PAL production was composed of 15g/L glucose,10g/L yeast extract,0.8g/L KH₂PO₄, 0.05g/L FeSO₄·7H₂O, 0.5g/L MgSO₄·7H₂O,5g/L NaCl,0.9g/L DL-phenylalanine,pH5.0.The cell was incubated at 30℃ with 5.0% seed culture on a shaker operated at 160r/min,harvested after 42h.The enzyme specific activity as high as 16.02mU/mg was obtained.The optimum transformation conditions were 13.0g/L trans-cinnamic acid,5.0mol/L ammonia,NH₃H₂O:NH₄HCO₃=4:3(v/w), a=2.5,0.03g/L cetylpyridinium bromide, pH10.5, 30℃,for 8 hours. The conversion ratio of cinnamic acid to phenylalanine was more than 42.36%,the accumulation of phenylalanine was 6.13g/L.