title: Continuous multistep synthesis of perillic acid from limonene by catalytic biofilms under segmented flow.
abstract: The efficiency of biocatalytic reactions involving industrially interesting reactants is often constrained by toxification of the applied biocatalyst. Here, we evaluated the combination of biologically and technologically inspired strategies to overcome toxicity-related issues during the multistep oxyfunctionalization of (R)-(+)-limonene to (R)-(+)-perillic acid. Pseudomonas putida GS1 catalyzing selective limonene oxidation via the p-cymene degradation pathway and recombinant Pseudomonas taiwanensis VLB120 were evaluated for continuous perillic acid production. A tubular segmented-flow biofilm reactor was used in order to relieve oxygen limitations and to enable membrane mediated substrate supply as well as efficient in situ product removal. Both P. putida GS1 and P. taiwanensis VLB120 developed a catalytic biofilm in this system. The productivity of wild-type P. putida GS1 encoding the enzymes for limonene bioconversion was highly dependent on the carbon source and reached 34 g Ltube-1day-1when glycerol was supplied. More than 10-fold lower productivities were reached irrespective of the applied carbon source when the recombinant P. taiwanensis VLB120 harboring p-cymene monooxygenase and p-cumic alcohol dehydrogenase was used as biocatalyst. The technical applicability for preparative perillic acid synthesis in the applied system was verified by purification of perillic acid from the outlet stream using an anion exchanger resin. This concept enabled the multistep production of perillic acid and which might be transferred to other reactions involving volatile reactants and toxic end-products. Biotechnol. Bioeng. 2017;114: 281-290. © 2016 Wiley Periodicals, Inc.
authors.affiliation: Department of Solar Materials, Helmholtz Centre for Environmental Research (UFZ), Permoserstrasse 15, 04318 Leipzig, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Willrodt
authors.affiliation: Department of Solar Materials, Helmholtz Centre for Environmental Research (UFZ), Permoserstrasse 15, 04318 Leipzig, Germany.
authors.forename: Babu
authors.initials: B
authors.lastname: Halan
authors.affiliation: Department of Biochemical and Chemical Engineering, Laboratory of Chemical Biotechnology, TU Dortmund University, Dortmund, Germany.
authors.forename: Lisa
authors.initials: L
authors.lastname: Karthaus
authors.affiliation: BRAIN Aktiengesellschaft, Zwingenberg, Germany.
authors.forename: Jessica
authors.initials: J
authors.lastname: Rehdorf
authors.affiliation: Department of Biochemical and Chemical Engineering, Laboratory of Chemical Biotechnology, TU Dortmund University, Dortmund, Germany.
authors.forename: Mattijs K
authors.initials: MK
authors.lastname: Julsing
authors.affiliation: Department of Solar Materials, Helmholtz Centre for Environmental Research (UFZ), Permoserstrasse 15, 04318 Leipzig, Germany.
authors.forename: Katja
authors.initials: K
authors.lastname: Buehler
authors.affiliation: Department of Solar Materials, Helmholtz Centre for Environmental Research (UFZ), Permoserstrasse 15, 04318 Leipzig, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Schmid
mesh_terms: D018441:Biofilms; D019149:Bioreactors / Q000382:microbiology; D003001:Cloning, Molecular; D053138:Cyclohexenes / Q000032:analysis / Q000302:isolation & purification / Q000378:metabolism*; D000077222:Limonene; D039821:Monoterpenes / Q000032:analysis / Q000302:isolation & purification / Q000378:metabolism*; D011549:Pseudomonas / Q000235:genetics / Q000378:metabolism; D016958:Pseudomonas putida / Q000201:enzymology / Q000235:genetics; D013729:Terpenes / Q000032:analysis / Q000378:metabolism*
chemical_list: D053138:Cyclohexenes; D039821:Monoterpenes; D013729:Terpenes; C078706:perillic acid; D000077222:Limonene
keywords: biofilm; continuous process; limonene; multistep oxyfunctionalization; perillic acid; segmented-flow