Synthesis, test and application of chirale fluorescence substrates to evaluate enzymatic processes in different reaction media

authored by: Torsten Knüttel, Hartmut Meyer, Thomas Scheper
Abstract: A pseudo-enantiomeric reaction was developed which was monitored on-line via 2D-fluorescence spectroscopy. During enzymatic reactions the enantiomeric excess can be followed on-line. Fluorescence spectroscopic detectable substrates, l-/d-phenylalanine-7-amido-4-methylcoumarine (l-/d-PheAMC) and l-/d-phenylalanine-7-amido-4-trifluoro-methylcoumarine (l-/d-PheAFC) were synthesized and deployed for applications in aqueous systems. Several proteases and esterases were tested to detect suitable biocatalysts for the enzymatic hydrolysis of the coumarine substrates. With α-chymotrypsin and the esterase from porcine liver, every coumarine substrate was hydrolyzed in an aqueous system and the determined enzymatic parameters were compared. The enantioselectivity and the enantiomeric excess of the enzymatic reactions were investigated. In simultaneous applications of the l- and d-substrates, the reactions were monitored on-line and the enantioselectivities, enantiomeric excesses and kinetic parameters were examined. To display the versatility of the on-line method, the enzymatic reactions were transferred to the organic solvent toluene. After the optimisation of the biocatalysis in toluene, the kinetic parameters were measured and the ee-parameters for the substrates and products were examined in off-line and on-line investigations.
Organisation(s): Institute of Technical Chemistry
External Organisation(s): Amersham Health
Type: Article
Journal: Enzyme and microbial technology
Volume: 37
Pages: 673-686
No. of pages: 14
ISSN: 0141-0229
Publication date: 14.06.2005
Publication status: Published
Peer reviewed: Yes
ASJC Scopus subject areas: Biotechnology, Bioengineering, Biochemistry, Applied Microbiology and Biotechnology
Electronic version(s): https://doi.org/10.1016/j.enzmictec.2004.04.026 (Access: Unknown)

BibTeX

@article{8f333c8c415c4409845630fd0dcecffa,
title = "Synthesis, test and application of chirale fluorescence substrates to evaluate enzymatic processes in different reaction media",
abstract = "A pseudo-enantiomeric reaction was developed which was monitored on-line via 2D-fluorescence spectroscopy. During enzymatic reactions the enantiomeric excess can be followed on-line. Fluorescence spectroscopic detectable substrates, l-/d-phenylalanine-7-amido-4-methylcoumarine (l-/d-PheAMC) and l-/d-phenylalanine-7-amido-4-trifluoro-methylcoumarine (l-/d-PheAFC) were synthesized and deployed for applications in aqueous systems. Several proteases and esterases were tested to detect suitable biocatalysts for the enzymatic hydrolysis of the coumarine substrates. With α-chymotrypsin and the esterase from porcine liver, every coumarine substrate was hydrolyzed in an aqueous system and the determined enzymatic parameters were compared. The enantioselectivity and the enantiomeric excess of the enzymatic reactions were investigated. In simultaneous applications of the l- and d-substrates, the reactions were monitored on-line and the enantioselectivities, enantiomeric excesses and kinetic parameters were examined. To display the versatility of the on-line method, the enzymatic reactions were transferred to the organic solvent toluene. After the optimisation of the biocatalysis in toluene, the kinetic parameters were measured and the ee-parameters for the substrates and products were examined in off-line and on-line investigations.",
keywords = "α-Chymotrypsin, 2D-fluorescence spectroscopy, Coumarines, Enantioselective hydrolysis, Esterase, On-line monitoring",
author = "Torsten Kn{\"u}ttel and Hartmut Meyer and Thomas Scheper",
note = "Funding information: The authors thank the Deutsche Forschungsgemeinschaft (DFG, as part of the Graduiertenkolleg) for financial support.",
year = "2005",
month = jun,
day = "14",
doi = "10.1016/j.enzmictec.2004.04.026",
language = "English",
volume = "37",
pages = "673--686",
journal = "Enzyme and microbial technology",
issn = "0141-0229",
publisher = "Elsevier Inc.",
number = "7",
}