為了以可再生的綠色生質化學品取代現今石油化學品，近年來許多相關的研究及製程陸續的發表與推出。目前尋找可再生的化合物中，2,5-furandicarboxylic acid (FDCA) 是未來具有發展性的化合物。其生產方式為由5-hydroxymethylfurfural（HMF）氧化得到中間產物5-(hydroxymethyl)furan-2-carboxylic acid (HMF acid)再繼續氧化成2,5-furandicarboxylic acid (FDCA)，此化合物可以替代聚合物PET生產過程中所需的對二甲酸。製造出的高分子材料對環境較PET更為友善。
本研究主要利用基因轉殖技術來產生出兩種不同酵素，分別為HMFH及HMFO，將HMFH及HMFO的基因序列分別接在載體上，利用可耐毒性的細菌Pseudomonas putida s12表達，並在Pseudomonas putida s12體內產生這兩種酵素，在高濃度的HMF下，利用Pseudomonas putida s12的全細胞做為生物催化劑，將HMF氧化並產生出中間產物HMF acid以及最終產物FDCA，並使HMFH和HMFO酵素結合，找出能使HMF 氧化成FDCA的最大量。目前實驗結果發現結合HMFH、HMFO和ALDH的效能最好，2小時即可產生FDCA，並濃度高達15mM，且HMF轉化成HMF acid的量高達約34mM，在4小時HMF完全氧化，並幾乎於24小時後中間產物HMF acid皆反應為最終產物FDCA。

Due to the indiscriminate use of fossil resources, the need for renewable feedstocks is become increase evident. 2,5-furandicarboxylic acid (FDCA) is a promising bio-based platform chemical. Production of FDCA needs to oxidize the 5-hydroxymethylfurfural（HMF）. When HMF is oxidized, it can become 5-(hydroxymethyl)furan-2-carboxylic acid (HMF acid), which is intermediate. Then HMF acid is oxidized and product the FDCA. FDCA is a green substitute for terephthalate in polyesters, and it is much friendlier to environmental.
In this study, using gene recombination to make two different enzyme, which are called HMFH and HMFO. Inserting the gene of HMFH and HMFO into the vector, then transformate the vector which includes gene of HMFH or HMFO into the bacteria Pseudomonas putida s12. When the solvent-tolerant Pseudomonas putida s12 express, the two enzyme HMFH and HMFO can produce in the bacteria. When add the high concentration of HMF (50mM) to the Pseudomonas putida s12 including HMFH or HMFO, the bacteria work as the whole-cell biocatalyst, HMF is oxidized to FDCA. We combined HMFH and HMFO together to find how to make the production of FDCA maximum. In this result, we found that combine HMFH,HMFO and ALDH have best result, at 2 hours, FDCA could be detected around 15mM
and HMF be converted to HMF acid ,which concentration of HMF acid is around 34mM. After 4 hours HMF disappeared, all be converted into HMF acid and FDCA; and after 24 hours HMF acid almost disappear, just only detected FDCA.

1. Kamm, B., Production of platform chemicals and synthesis gas from biomass. Angew Chem Int Ed Engl, 2007. 46(27): p. 5056-8.
2. T.Werpy and G.Petersen, <Top Value Added Chemicals from Biomass Volume I—Results of Screening for Potential Candidates from Sugars and Synthesis Gas.pdf>. 2004. 1: p. 26-28.
3. Carlini, C., et al., Selective oxidation of 5-hydroxymethyl-2-furaldehyde to furan-2,5-dicarboxaldehyde by catalytic systems based on vanadyl phosphate. Applied Catalysis A: General, 2005. 289(2): p. 197-204.
4. Koopman, F., et al., Efficient whole-cell biotransformation of 5-(hydroxymethyl)furfural into FDCA, 2,5-furandicarboxylic acid. Bioresour Technol, 2010. 101(16): p. 6291-6.
5. Koopman, F., et al., Identification and characterization of the furfural and 5-(hydroxymethyl)furfural degradation pathways of Cupriavidus basilensis HMF14. Proc Natl Acad Sci U S A, 2010. 107(11): p. 4919-24.
6. Dijkman, W.P. and M.W. Fraaije, Discovery and Characterization of a 5-Hydroxymethylfurfural Oxidase from Methylovorus sp. Strain MP688. ASM journal, 2014. 80: p. 1082-1090.
7. Zhang, J., et al., Advances in catalytic production of bio-based polyester monomer 2,5-furandicarboxylic acid derived from lignocellulosic biomass. Carbohydr Polym, 2015. 130: p. 420-8.
8. Dijkman, W.P., D.E. Groothuis, and M.W. Fraaije, Enzyme-catalyzed oxidation of 5-hydroxymethylfurfural to furan-2,5-dicarboxylic acid. Angew Chem Int Ed Engl, 2014. 53(25): p. 6515-8.
9. Siankevich, S., et al., A novel platinum nanocatalyst for the oxidation of 5-Hydroxymethylfurfural into 2,5-Furandicarboxylic acid under mild conditions. Journal of Catalysis, 2014. 315: p. 67-74.
10. Yi, G., et al., Purification of biomass-derived 5-hydroxymethylfurfural and its catalytic conversion to 2,5-furandicarboxylic Acid. ChemSusChem, 2014. 7(8): p. 2131-5.
11. Archana Jain, S.C.J., Kandalam V.Ramanujachary, Amos Mugweru, Selective oxidation of 5-hydroxymethyl-2-furfural to furan-2,5-dicarboxylic acid over spinel mixed metal oxide catalyst. catalysis Communication, 2015: p. 179-182.
12. Petinakis, E., et al., Natural Fibre Bio-Composites Incorporating Poly(Lactic Acid). 2013.
13. Tan, G.-Y., et al., Start a Research on Biopolymer Polyhydroxyalkanoate (PHA): A Review. Polymers, 2014. 6(3): p. 706-754.
14. Japu, C., et al., Bio-based poly(ethylene terephthalate) copolyesters made from cyclic monomers derived from tartaric acid. Polymer, 2014. 55(10): p. 2294-2304.
15. Max M. Houck, R., E.Menold, Rebecca A. Huff, POLY(TRIMETHYLENET EREPHTHALATE)A“NEW”TYPE OF POLYESTER FIBRE. prombles of Forensic Science, 2001. XLVI: p. 217-221.
16. A.M. Radder, H.L., C.A. van Blitterswijk, Bone-bonding behaviour of poly(ethylene oxide) - polybutylene terephthalate copolymer coatings and bulk implants a comparative study. Biomaterials 1995. 16(7): p. 507-513.
17. Wierckx, N., et al., Isolation and characterization of Cupriavidus basilensis HMF14 for biological removal of inhibitors from lignocellulosic hydrolysate. Microb Biotechnol, 2010. 3(3): p. 336-43.
18. Wery, J., et al., An insertion sequence prepares Pseudomonas putida S12 for severe solvent stress. J Biol Chem, 2001. 276(8): p. 5700-6.
19. Foti, M., R. Medici, and H.J. Ruijssenaars, Biological production of monoethanolamine by engineered Pseudomonas putida S12. J Biotechnol, 2013. 167(3): p. 344-9.
20. Fei Tao, Y.L., Qian Luo, Fei Su, Youqiang Xu, Fuli Li, Cuiqing Ma, Ping Xu, Novel organic solvent-responsive expression vectors for biocatalysis Application for development of an organic solvent-tolerant biodesulfurizing strain. Bioresour Technology, 2011. 102: p. 9380-9387.
21. Jan Wery, J.A.M.d.B., SOLVENT-TOLERANCE OF PSEUDOMONADS A NEW DEGREE OF FREEDOM IN BIOCATALYSIS. Kluwer Academic/Plenum Publishers, New York, 2004.
22. Nijkamp, K., et al., The solvent-tolerant Pseudomonas putida S12 as host for the production of cinnamic acid from glucose. Appl Microbiol Biotechnol, 2005. 69(2): p. 170-7.
23. prather, K., Integrated Chemical Engineering (ICE) Topics: Biocatalysis Lecture #5 – Using Whole Cells as Biocatalysts Why When, Growth vs Conversion (Screening). 2004.
24. Ishige, T., K. Honda, and S. Shimizu, Whole organism biocatalysis. Curr Opin Chem Biol, 2005. 9(2): p. 174-80.
25. Dueber, J.E., et al., Synthetic protein scaffolds provide modular control over metabolic flux. Nat Biotechnol, 2009. 27(8): p. 753-9.
26. Jeon, S.D., et al., Analysis of selective, high protein-protein binding interaction of cohesin-dockerin complex using biosensing methods. Biosens Bioelectron, 2012. 35(1): p. 382-9.
27. Jeon, S.D., et al., An enhanced protein-protein interaction based on enzymatic complex through replacement of the recognition site. Int J Biol Macromol, 2015. 75: p. 1-6.
28. Van Dung Phama , S.H.L., Si Jae Parkc, Soon Ho Honga, Gamma-aminobutyric acid production through GABA shunt by synthetic scaffolds introduction in recombinant Escherichia coli. Biochemical Engineering Journal, 2015.
29. Moon, T.S., et al., Use of modular, synthetic scaffolds for improved production of glucaric acid in engineered E. coli. Metab Eng, 2010. 12(3): p. 298-305.
30. Wang, Y. and O. Yu, Synthetic scaffolds increased resveratrol biosynthesis in engineered yeast cells. Journal of Biotechnology, 2012: p. 258-260.
31. S. HARTMANS, J.P.S., t M. J. VAN DER WERF, F. VOLKERING, AND J. A. M. DE BONT, Metabolism of Styrene Oxide and 2-Phenylethanol in the Styrene-Degrading Xanthobacter Strain 124X. Applied and Environment Microbiology, 1989. 55(11): p. 2859-2855.
32. Pool, H., et al., Novel high-level constitutive expression system, pHCE vector, for aconvenient and cost-effective soluble production of human tumor necrosisfactor-α.pdf. Kluwer academic publishers, 2002: p. 1185-1189.