Florin Irimie | Babes-Bolyai University (original) (raw)

Papers by Florin Irimie

Environmental engineering and management journal

We have previously described the molecular cloning and expression in an Escherichia coli system a... more We have previously described the molecular cloning and expression in an Escherichia coli system and the characterization of a thermostable esterase/lipase from Anoxybacillus flavithermus T1. We herein report on the optim ization of the expression process. When using isopropyl--D-thiogalactopyranoside (IPTG) for induction, the high est protein yield was obtained at 30°C, with 0.4 mM IPTG and 1 h induction time. Similar results were obtained at 37°C, but with a higher IPTG concentration (5 mM) and after 8h of induction, which makes the former a better option in terms of cost and time-efectiveness. Better results yet were attained with lactose, a very attractive option, given its high availability, low cost and low toxicity to the host cells. Recovery of the active enzyme from the periplasmic space was highest with a lysis buffer which combines osmotic shock with a membrane destabilization effect (Tris-sucrose/EDTA 1 mM/MgSO4 5 mM). The freeze-thaw treatment yielded similar results, w...

RSC Advances

The EKR of some heteroaromatic secondary ethanols with tailored sol–gel immobilized lipases in ba... more The EKR of some heteroaromatic secondary ethanols with tailored sol–gel immobilized lipases in batch and continuous-flow reactors was studied. The productivity in continuous-flow mode is higher than in batch mode.

Reaction Chemistry & Engineering

A new, efficient lipase-mediated kinetic resolution–click-reaction-based procedure is presented f... more A new, efficient lipase-mediated kinetic resolution–click-reaction-based procedure is presented for the production of both enantiomers of various 1-(hetero)aromatic ethanols.

Environmental Engineering and Management Journal

We have previously described the molecular cloning and expression in an Escherichia coli system a... more We have previously described the molecular cloning and expression in an Escherichia coli system and the characterization of a thermostable esterase/lipase from Anoxybacillus flavithermus T1. We herein report on the optimization of the expression process. When using isopropyl--D-thiogalactopyranoside (IPTG) for induction, the highest protein yield was obtained at 30°C, with 0.4 mM IPTG and 1 h induction time. Similar results were obtained at 37°C, but with a higher IPTG concentration (5 mM) and after 8h of induction, which makes the former a better option in terms of cost and time-efectiveness. Better results yet were attained with lactose, a very attractive option, given its high availability, low cost and low toxicity to the host cells. Recovery of the active enzyme from the periplasmic space was highest with a lysis buffer which combines osmotic shock with a membrane destabilization effect (Tris-sucrose/EDTA 1 mM/MgSO 4 5 mM). The freeze-thaw treatment yielded similar results, while treatment with 1% organic solvent (chloroform or DMSO), while effective in permeabilization of the cell membrane, exerted a certain inhibitory effect upon the enzyme.

Molecules

The Amano lipase from Pseudomonas fluorescens (L-AK) was covalently immobilized on various carbon... more The Amano lipase from Pseudomonas fluorescens (L-AK) was covalently immobilized on various carbon nanomaterials (functionalized single-walled carbon nanotubes and graphene oxide) and tested for biodiesel production. Using the most active lipase preparation (covalently immobilized L-AK on SwCNTNH2 derivatized with glycerol diglycidyl ether) under optimal conditions, quasi-complete conversion (>99%) of sunflower oil was obtained after only 4 h reaction time. Moreover, the biocatalyst maintained more than 99% of its initial activity in the batch system after multiple recycling experiments.

Molecules

Lipase B from Candida antarctica immobilized by covalent binding on sebacoyl-activated chitosan-c... more Lipase B from Candida antarctica immobilized by covalent binding on sebacoyl-activated chitosan-coated magnetic nanoparticles proved to be an efficient biocatalyst (49.2–50% conversion in 3–16 h and >96% enantiomeric excess) for the enzymatic kinetic resolution of some racemic heteroarylethanols through transesterification with vinyl acetate. Under optimal conditions (vinyl acetate, n-hexane, 45 °C), the biocatalyst remains active after 10 cycles.

Studia Universitatis Babeș Bolyai Chemia, 2003

Molecules, 2015

In this paper we describe the chemoenzymatic synthesis of enantiopure L-2-arylthiazol-4-yl alanin... more In this paper we describe the chemoenzymatic synthesis of enantiopure L-2-arylthiazol-4-yl alanines starting from their racemic N-acetyl derivatives; by combining the lipase-catalysed dynamic kinetic resolution of oxazol-5(4H)-ones with a chemical and an enzymatic enantioselective hydrolytic step affording the desired products in good yields (74%-78%) and high enantiopurities (ee > 99%). The developed procedure exploits the utility of the single-walled carbon nanotubes-bound diethylaminoethanol as mild and efficient racemisation agent for the dynamic kinetic resolution of the corresponding oxazolones.

Tetrahedron: Asymmetry, 2015

ABSTRACT

Angewandte Chemie (International ed. in English), Jan 27, 2015

Molecules, 2015

This paper describes the biocatalytic synthesis of new Mannich bases containing various heterocyc... more This paper describes the biocatalytic synthesis of new Mannich bases containing various heterocyclic rings (thiazole, furane, thiophene, pyridine) by applying the lipase catalyzed trimolecular condensation of the corresponding heterocyclic aldehydes with acetone and primary aromatic amines, in mild and eco-friendly reaction conditions. The obtained Mannich bases were acylated to their corresponding N-acetyl derivatives. All compounds were characterized by 1 H-NMR, 13 C-NMR and MS spectrometry.

Angewandte Chemie, 2014

Cation-π interactions to cognate ligands in enzymes have key roles in ligand binding and enzymati... more Cation-π interactions to cognate ligands in enzymes have key roles in ligand binding and enzymatic catalysis. We have deciphered the key functional role of both charged and aromatic residues within the choline binding subsite of CTP:phosphocholine cytidylyltransferase and choline kinase from Plasmodium falciparum. Comparison of quaternary ammonium binding site structures revealed a general composite aromatic box pattern of enzyme recognition sites, well distinguished from the aromatic box recognition site of receptors.

Tetrahedron: Asymmetry, 2014

ABSTRACT Both enantiomers of bufuralol are pharmaceutically important molecules. While the (S)-is... more ABSTRACT Both enantiomers of bufuralol are pharmaceutically important molecules. While the (S)-isomer with a higher β-blocking activity is recommended for hypertension treatment, the (R)-enantiomer can be used as marker of hepatic activity. In this paper two new alternative approaches are described for their chemo-enzymatic synthesis, providing both highly enantiomerically enriched stereoisomers of the target molecule (ee 96–98%). One route is based on the baker’s yeast mediated stereoselective biotransformation of α-substituted ketones, and the other one on the lipase mediated kinetic resolution of the racemic bromoethanol.

ChemCatChem, 2013

ABSTRACT Racemic nitrophenylalanines and (E)‐nitrophenylacrylates are synthesized from the corres... more ABSTRACT Racemic nitrophenylalanines and (E)‐nitrophenylacrylates are synthesized from the corresponding aldehydes. Both products are important for the examination of the mechanism of action of phenylalanine ammonia lyase (PAL). For the reaction of the rac‐nitrophenylalanines with both wild type (wt) PAL and an 4‐methylideneimidazole‐5‐one (MIO)‐less mutant, the kinetic constants K m and V max are determined and compared with those of the natural substrate L‐phenylalanine: the K m values for the racemic nitrophenylalanines with wt PAL are up to 9 times higher, however, the V max values are up to 5 times lower. Compared to wt PAL, the catalytic activity of MIO‐less PAL mutant for the deamination of L‐phenylalanine is approximately 400 times, while that for 3‐nitrophenylalanine is approximately 50 times lower. Both wt and MIO‐less PALs are enantioselective for L‐nitrophenylalanines. Thus, enantiopure D‐nitrophenylalanines can be biosynthesized from racemic substrates. The biocatalytic synthesis of the corresponding L‐enantiomers is achieved by the reverse reaction, starting from (E)‐nitrophenylacrylates. Both enantiomeric products obtained with wt and MIO‐less PAL are spectroscopically and chromatographically characterized and their optical rotations measured.

Tetrahedron: Asymmetry, 2012

Starting from the racemic 2-benzofuranyl-and 2-benzo[b]thiophenyl-2-hydroxyacetic acid ethyl este... more Starting from the racemic 2-benzofuranyl-and 2-benzo[b]thiophenyl-2-hydroxyacetic acid ethyl esters as substrates, a general method was developed for the efficient synthesis of the corresponding highly enantiomerically enriched (ee up to 99%) (R)-and (S)-2-heteroaryl-2-hydroxyacetic acids.

Environmental engineering and management journal

We have previously described the molecular cloning and expression in an Escherichia coli system a... more We have previously described the molecular cloning and expression in an Escherichia coli system and the characterization of a thermostable esterase/lipase from Anoxybacillus flavithermus T1. We herein report on the optim ization of the expression process. When using isopropyl--D-thiogalactopyranoside (IPTG) for induction, the high est protein yield was obtained at 30°C, with 0.4 mM IPTG and 1 h induction time. Similar results were obtained at 37°C, but with a higher IPTG concentration (5 mM) and after 8h of induction, which makes the former a better option in terms of cost and time-efectiveness. Better results yet were attained with lactose, a very attractive option, given its high availability, low cost and low toxicity to the host cells. Recovery of the active enzyme from the periplasmic space was highest with a lysis buffer which combines osmotic shock with a membrane destabilization effect (Tris-sucrose/EDTA 1 mM/MgSO4 5 mM). The freeze-thaw treatment yielded similar results, w...

RSC Advances

The EKR of some heteroaromatic secondary ethanols with tailored sol–gel immobilized lipases in ba... more The EKR of some heteroaromatic secondary ethanols with tailored sol–gel immobilized lipases in batch and continuous-flow reactors was studied. The productivity in continuous-flow mode is higher than in batch mode.

Reaction Chemistry & Engineering

A new, efficient lipase-mediated kinetic resolution–click-reaction-based procedure is presented f... more A new, efficient lipase-mediated kinetic resolution–click-reaction-based procedure is presented for the production of both enantiomers of various 1-(hetero)aromatic ethanols.

Environmental Engineering and Management Journal

We have previously described the molecular cloning and expression in an Escherichia coli system a... more We have previously described the molecular cloning and expression in an Escherichia coli system and the characterization of a thermostable esterase/lipase from Anoxybacillus flavithermus T1. We herein report on the optimization of the expression process. When using isopropyl--D-thiogalactopyranoside (IPTG) for induction, the highest protein yield was obtained at 30°C, with 0.4 mM IPTG and 1 h induction time. Similar results were obtained at 37°C, but with a higher IPTG concentration (5 mM) and after 8h of induction, which makes the former a better option in terms of cost and time-efectiveness. Better results yet were attained with lactose, a very attractive option, given its high availability, low cost and low toxicity to the host cells. Recovery of the active enzyme from the periplasmic space was highest with a lysis buffer which combines osmotic shock with a membrane destabilization effect (Tris-sucrose/EDTA 1 mM/MgSO 4 5 mM). The freeze-thaw treatment yielded similar results, while treatment with 1% organic solvent (chloroform or DMSO), while effective in permeabilization of the cell membrane, exerted a certain inhibitory effect upon the enzyme.

Molecules

The Amano lipase from Pseudomonas fluorescens (L-AK) was covalently immobilized on various carbon... more The Amano lipase from Pseudomonas fluorescens (L-AK) was covalently immobilized on various carbon nanomaterials (functionalized single-walled carbon nanotubes and graphene oxide) and tested for biodiesel production. Using the most active lipase preparation (covalently immobilized L-AK on SwCNTNH2 derivatized with glycerol diglycidyl ether) under optimal conditions, quasi-complete conversion (>99%) of sunflower oil was obtained after only 4 h reaction time. Moreover, the biocatalyst maintained more than 99% of its initial activity in the batch system after multiple recycling experiments.

Molecules

Lipase B from Candida antarctica immobilized by covalent binding on sebacoyl-activated chitosan-c... more Lipase B from Candida antarctica immobilized by covalent binding on sebacoyl-activated chitosan-coated magnetic nanoparticles proved to be an efficient biocatalyst (49.2–50% conversion in 3–16 h and >96% enantiomeric excess) for the enzymatic kinetic resolution of some racemic heteroarylethanols through transesterification with vinyl acetate. Under optimal conditions (vinyl acetate, n-hexane, 45 °C), the biocatalyst remains active after 10 cycles.

Studia Universitatis Babeș Bolyai Chemia, 2003

Molecules, 2015

In this paper we describe the chemoenzymatic synthesis of enantiopure L-2-arylthiazol-4-yl alanin... more In this paper we describe the chemoenzymatic synthesis of enantiopure L-2-arylthiazol-4-yl alanines starting from their racemic N-acetyl derivatives; by combining the lipase-catalysed dynamic kinetic resolution of oxazol-5(4H)-ones with a chemical and an enzymatic enantioselective hydrolytic step affording the desired products in good yields (74%-78%) and high enantiopurities (ee > 99%). The developed procedure exploits the utility of the single-walled carbon nanotubes-bound diethylaminoethanol as mild and efficient racemisation agent for the dynamic kinetic resolution of the corresponding oxazolones.

Tetrahedron: Asymmetry, 2015

ABSTRACT

Angewandte Chemie (International ed. in English), Jan 27, 2015

Molecules, 2015

This paper describes the biocatalytic synthesis of new Mannich bases containing various heterocyc... more This paper describes the biocatalytic synthesis of new Mannich bases containing various heterocyclic rings (thiazole, furane, thiophene, pyridine) by applying the lipase catalyzed trimolecular condensation of the corresponding heterocyclic aldehydes with acetone and primary aromatic amines, in mild and eco-friendly reaction conditions. The obtained Mannich bases were acylated to their corresponding N-acetyl derivatives. All compounds were characterized by 1 H-NMR, 13 C-NMR and MS spectrometry.

Angewandte Chemie, 2014

Cation-π interactions to cognate ligands in enzymes have key roles in ligand binding and enzymati... more Cation-π interactions to cognate ligands in enzymes have key roles in ligand binding and enzymatic catalysis. We have deciphered the key functional role of both charged and aromatic residues within the choline binding subsite of CTP:phosphocholine cytidylyltransferase and choline kinase from Plasmodium falciparum. Comparison of quaternary ammonium binding site structures revealed a general composite aromatic box pattern of enzyme recognition sites, well distinguished from the aromatic box recognition site of receptors.

Tetrahedron: Asymmetry, 2014

ABSTRACT Both enantiomers of bufuralol are pharmaceutically important molecules. While the (S)-is... more ABSTRACT Both enantiomers of bufuralol are pharmaceutically important molecules. While the (S)-isomer with a higher β-blocking activity is recommended for hypertension treatment, the (R)-enantiomer can be used as marker of hepatic activity. In this paper two new alternative approaches are described for their chemo-enzymatic synthesis, providing both highly enantiomerically enriched stereoisomers of the target molecule (ee 96–98%). One route is based on the baker’s yeast mediated stereoselective biotransformation of α-substituted ketones, and the other one on the lipase mediated kinetic resolution of the racemic bromoethanol.

ChemCatChem, 2013

ABSTRACT Racemic nitrophenylalanines and (E)‐nitrophenylacrylates are synthesized from the corres... more ABSTRACT Racemic nitrophenylalanines and (E)‐nitrophenylacrylates are synthesized from the corresponding aldehydes. Both products are important for the examination of the mechanism of action of phenylalanine ammonia lyase (PAL). For the reaction of the rac‐nitrophenylalanines with both wild type (wt) PAL and an 4‐methylideneimidazole‐5‐one (MIO)‐less mutant, the kinetic constants K m and V max are determined and compared with those of the natural substrate L‐phenylalanine: the K m values for the racemic nitrophenylalanines with wt PAL are up to 9 times higher, however, the V max values are up to 5 times lower. Compared to wt PAL, the catalytic activity of MIO‐less PAL mutant for the deamination of L‐phenylalanine is approximately 400 times, while that for 3‐nitrophenylalanine is approximately 50 times lower. Both wt and MIO‐less PALs are enantioselective for L‐nitrophenylalanines. Thus, enantiopure D‐nitrophenylalanines can be biosynthesized from racemic substrates. The biocatalytic synthesis of the corresponding L‐enantiomers is achieved by the reverse reaction, starting from (E)‐nitrophenylacrylates. Both enantiomeric products obtained with wt and MIO‐less PAL are spectroscopically and chromatographically characterized and their optical rotations measured.

Tetrahedron: Asymmetry, 2012

Starting from the racemic 2-benzofuranyl-and 2-benzo[b]thiophenyl-2-hydroxyacetic acid ethyl este... more Starting from the racemic 2-benzofuranyl-and 2-benzo[b]thiophenyl-2-hydroxyacetic acid ethyl esters as substrates, a general method was developed for the efficient synthesis of the corresponding highly enantiomerically enriched (ee up to 99%) (R)-and (S)-2-heteroaryl-2-hydroxyacetic acids.