As a clinical dose requires a minimum of 106 cells per kilogram of patients, it is, therefore, cr... more As a clinical dose requires a minimum of 106 cells per kilogram of patients, it is, therefore, crucial to develop a scalable method of production of Wharton Jelly mesenchymal stem cells (WJ‐MSCs) with maintained inner characteristics. Scalable expansion of WJ‐MSCs on microcarriers usually found in cell culture, involves specific cell detachment using trypsin and could have harmful effects on cells. In this study, the performance of batch, fed‐batch, and perfused‐continuous mode of culture were compared. The batch and fed‐batch modes resulted in expansion factors of 5 and 43, respectively. The perfused‐continuous mode strategy consisted of the implementation of a settling tube inside the bioreactor. The diameter of the tube was calculated to maintain microcarriers colonized by cells in the bioreactor whereas empty microcarriers (responsible for potentially damaging collisions) were removed, using a continuous flow rate based on MSCs physiological requirements. Thanks to this strategy, a maximal number of 800 million cells was obtained in a 1.5 L bioreactor in 10 days. Lastly, online dielectric spectroscopy was implemented in the bioreactor and indicated that cell growth could be monitored during the culture.
Abstract Aminoacylases from a crude extract from Streptomyces ambofaciens were described as attra... more Abstract Aminoacylases from a crude extract from Streptomyces ambofaciens were described as attractive way for the synthesis of amino acids biobased surfactants. The aim of this study is to investigate the influence of the porosity and the APTES functionalization of the silica support on the immobilization and the activities of these aminoacylases. To reach this goal, a mesoporous (SBA-15) and of meso-macroporous (MMS) silica material without and with functionalization with APTES were used. The results show that the use of non-functionalized supports led to a poor enzyme loading due to electrostatic repulsion between the enzyme and the silica support. The APTES functionalization of the silica materials allow a better enzyme loading by favouring enzyme-support interactions with an increase of the immobilization rate by increasing the APTES functionalization rate from 0 to 10% especially when using MMS. However, the measurement of the catalytic performances of the enzymes immobilized on the materials shows a significant improvement of the conversion rate by increasing the APTES percentage in the case of SBA-15 unlike the enzymes immobilized on the functionalized MMS. This behaviour can be explained by the better substrate accessibility to the enzyme immobilized on the SBA-15-based materials which is mainly located on the external surface area of the functionalized materials. The cross linking of the aminoacylases by using glutaraldehyde led to a loss of the activity due to enzyme inactivation. The LC-MS-MS analysis of the product of lysine acylation reaction shows that the regioselectivity of the enzymes after immobilization is not modified.
The presence of aminoacylase activities was investigated in a crude extract of Streptomyces ambof... more The presence of aminoacylase activities was investigated in a crude extract of Streptomyces ambofaciens ATCC23877. First activities catalyzing the hydrolysis of N‐α or ε‐acetyl‐L‐lysine were identified. Furthermore, the acylation of lysine and different peptides was studied and compared with results obtained with lipase B of Candida antarctica (CALB). Different regioselectivities were demonstrated for the two classes of enzymes. CALB was able to catalyze acylation only on the ε‐position whereas the crude extract from S. ambofaciens possessed the rare ability to catalyze the N‐acylation on the α‐position of the lysine or of the amino‐acid in N‐terminal position of peptides. Two genes, SAM23877_1485 and SAM23877_1734, were identified in the genome of Streptomyces ambofaciens ATCC23877 whose products show similarities with the previously identified aminoacylases from Streptomyces mobaraensis. The proteins encoded by these two genes were responsible for the major aminoacylase hydrolytic...
The aim of the present work was to obtain microbial lipids (single-cell oils and SCOs) from oleag... more The aim of the present work was to obtain microbial lipids (single-cell oils and SCOs) from oleaginous yeast cultivated on biodiesel-derived glycerol and subsequently proceed to the enzymatic synthesis of high-value biosurfactant-type molecules in an aqueous medium, with SCOs implicated as acyl donors (ADs). Indeed, the initial screening of five non-conventional oleaginous yeasts revealed that the most important lipid producer was the microorganism Cryptococcus curvatus ATCC 20509. SCO production was optimised according to the nature of the nitrogen source and the initial concentration of glycerol (Glyc0) employed in the medium. Lipids up to 50% w/w in dry cell weight (DCW) (SCOmax = 6.1 g/L) occurred at Glyc0 ≈ 70 g/L (C/N ≈ 80 moles/moles). Thereafter, lipids were recovered and were subsequently used as ADs in the N-acylation reaction catalysed by aminoacylases produced from Streptomyces ambofaciens ATCC 23877 under aqueous conditions, while Candida antarctica lipase B (CALB) was ...
The ferulic acid (FA)-oxidation by Myceliophthora thermophila laccase was performed in phosphate ... more The ferulic acid (FA)-oxidation by Myceliophthora thermophila laccase was performed in phosphate buffer at 30 °C and pH 7.5 as an eco-friendly procedure. LC-MS analysis showed that oxidation products were four dehydrodimers (P1, P2, P3, P5) at MM = 386 g/mol, two dehydrotetramers (P6, P7) at MM = 770 g/mol and one decarboxylated dehydrodimer (P4) at MM = 340 g/mol. Structural characterization showed that FA-dehydrodimers were symmetric for P1 and P5 while asymmetric for P2, P3 and P4. Physicochemical characterization showed that oxidation products presented a higher lipophilicity than that of FA. Moreover, symmetric dimers and tetra dimers had a higher melting point compared to FA and its asymmetric dimers. Antioxidant and anti-proliferative assessments indicated that enzymatic oligomerization increased antioxidant and anti-proliferative properties of oxidation products for P2, P3 and P6 compared to FA. Finally, this enzymatic process in water could produce new molecules, having goo...
A new glyco-phenol was produced by the coupling between glucosamine (Glu) and ferulic acid (FA) u... more A new glyco-phenol was produced by the coupling between glucosamine (Glu) and ferulic acid (FA) using Myceliophthora thermophila laccase as biocatalyst in mild conditions (distilled water and 30°C) as an environmentally friendly process. Results indicated that the enzymatic reaction created a new derivative (FA-Glu), produced from coupling between Glu and FA by covalent bonds. By the high production of (FA-Glu) derivative and its stability, the optimal ratio of (FA: Glu) was of (1:1) at optimal time reaction of 6 h. Under these optimal conditions, almost 55% of -NH2 groups on Glu were bound with FA-oxidation products. The new derivative showed higher hydrophobic character than Glu due to the presence of FA in its structure. LC-MS analysis showed that (FA-Glu) derivative exhibited a molecular mass at MM 713 g/mol containing one Glu-molecule and three FA-molecules after decarboxylation. Furthermore, the new derivative presented good antioxidant and anti-proliferative activities in comparison with Glu and FA. These results suggest that the enzymatic conjugation between Glu and FA is a promising process to produce a new glyco-phenol having good functional properties for potential applications. This article is protected by copyright. All rights reserved.
Many studies underline the great benefits of yeast extracts (YE), used as supplements in animal f... more Many studies underline the great benefits of yeast extracts (YE), used as supplements in animal free culture media, on cell growth and recombinant protein production [1,2]. Nevertheless, their unknown composition and batch-to-batch variability of commercial YE remain constraints for industrial processes [3,4]. Consequently, the identification of bioactive YE molecules is challenging for process reliability. The main strategy to respond upon this problem is to fractionate the extract and then to characterize the fractions. So far, several fractionation processes such as ultrafiltration [5], gel filtration chromatography [6] or alcoholic precipitation [7] have been investigated. These studies suggested that the active components were mainly small molecules (< 1000 Da). But, the other physico-chemical properties of YE components have been poorly studied until now. In regards of this statement, it is proposed to get better knowledge of the YE molecules leading to an improvement of CH...
All in-text references underlined in blue are linked to publications on ResearchGate, letting you... more All in-text references underlined in blue are linked to publications on ResearchGate, letting you access and read them immediately.
As a clinical dose requires a minimum of 106 cells per kilogram of patients, it is, therefore, cr... more As a clinical dose requires a minimum of 106 cells per kilogram of patients, it is, therefore, crucial to develop a scalable method of production of Wharton Jelly mesenchymal stem cells (WJ‐MSCs) with maintained inner characteristics. Scalable expansion of WJ‐MSCs on microcarriers usually found in cell culture, involves specific cell detachment using trypsin and could have harmful effects on cells. In this study, the performance of batch, fed‐batch, and perfused‐continuous mode of culture were compared. The batch and fed‐batch modes resulted in expansion factors of 5 and 43, respectively. The perfused‐continuous mode strategy consisted of the implementation of a settling tube inside the bioreactor. The diameter of the tube was calculated to maintain microcarriers colonized by cells in the bioreactor whereas empty microcarriers (responsible for potentially damaging collisions) were removed, using a continuous flow rate based on MSCs physiological requirements. Thanks to this strategy, a maximal number of 800 million cells was obtained in a 1.5 L bioreactor in 10 days. Lastly, online dielectric spectroscopy was implemented in the bioreactor and indicated that cell growth could be monitored during the culture.
Abstract Aminoacylases from a crude extract from Streptomyces ambofaciens were described as attra... more Abstract Aminoacylases from a crude extract from Streptomyces ambofaciens were described as attractive way for the synthesis of amino acids biobased surfactants. The aim of this study is to investigate the influence of the porosity and the APTES functionalization of the silica support on the immobilization and the activities of these aminoacylases. To reach this goal, a mesoporous (SBA-15) and of meso-macroporous (MMS) silica material without and with functionalization with APTES were used. The results show that the use of non-functionalized supports led to a poor enzyme loading due to electrostatic repulsion between the enzyme and the silica support. The APTES functionalization of the silica materials allow a better enzyme loading by favouring enzyme-support interactions with an increase of the immobilization rate by increasing the APTES functionalization rate from 0 to 10% especially when using MMS. However, the measurement of the catalytic performances of the enzymes immobilized on the materials shows a significant improvement of the conversion rate by increasing the APTES percentage in the case of SBA-15 unlike the enzymes immobilized on the functionalized MMS. This behaviour can be explained by the better substrate accessibility to the enzyme immobilized on the SBA-15-based materials which is mainly located on the external surface area of the functionalized materials. The cross linking of the aminoacylases by using glutaraldehyde led to a loss of the activity due to enzyme inactivation. The LC-MS-MS analysis of the product of lysine acylation reaction shows that the regioselectivity of the enzymes after immobilization is not modified.
The presence of aminoacylase activities was investigated in a crude extract of Streptomyces ambof... more The presence of aminoacylase activities was investigated in a crude extract of Streptomyces ambofaciens ATCC23877. First activities catalyzing the hydrolysis of N‐α or ε‐acetyl‐L‐lysine were identified. Furthermore, the acylation of lysine and different peptides was studied and compared with results obtained with lipase B of Candida antarctica (CALB). Different regioselectivities were demonstrated for the two classes of enzymes. CALB was able to catalyze acylation only on the ε‐position whereas the crude extract from S. ambofaciens possessed the rare ability to catalyze the N‐acylation on the α‐position of the lysine or of the amino‐acid in N‐terminal position of peptides. Two genes, SAM23877_1485 and SAM23877_1734, were identified in the genome of Streptomyces ambofaciens ATCC23877 whose products show similarities with the previously identified aminoacylases from Streptomyces mobaraensis. The proteins encoded by these two genes were responsible for the major aminoacylase hydrolytic...
The aim of the present work was to obtain microbial lipids (single-cell oils and SCOs) from oleag... more The aim of the present work was to obtain microbial lipids (single-cell oils and SCOs) from oleaginous yeast cultivated on biodiesel-derived glycerol and subsequently proceed to the enzymatic synthesis of high-value biosurfactant-type molecules in an aqueous medium, with SCOs implicated as acyl donors (ADs). Indeed, the initial screening of five non-conventional oleaginous yeasts revealed that the most important lipid producer was the microorganism Cryptococcus curvatus ATCC 20509. SCO production was optimised according to the nature of the nitrogen source and the initial concentration of glycerol (Glyc0) employed in the medium. Lipids up to 50% w/w in dry cell weight (DCW) (SCOmax = 6.1 g/L) occurred at Glyc0 ≈ 70 g/L (C/N ≈ 80 moles/moles). Thereafter, lipids were recovered and were subsequently used as ADs in the N-acylation reaction catalysed by aminoacylases produced from Streptomyces ambofaciens ATCC 23877 under aqueous conditions, while Candida antarctica lipase B (CALB) was ...
The ferulic acid (FA)-oxidation by Myceliophthora thermophila laccase was performed in phosphate ... more The ferulic acid (FA)-oxidation by Myceliophthora thermophila laccase was performed in phosphate buffer at 30 °C and pH 7.5 as an eco-friendly procedure. LC-MS analysis showed that oxidation products were four dehydrodimers (P1, P2, P3, P5) at MM = 386 g/mol, two dehydrotetramers (P6, P7) at MM = 770 g/mol and one decarboxylated dehydrodimer (P4) at MM = 340 g/mol. Structural characterization showed that FA-dehydrodimers were symmetric for P1 and P5 while asymmetric for P2, P3 and P4. Physicochemical characterization showed that oxidation products presented a higher lipophilicity than that of FA. Moreover, symmetric dimers and tetra dimers had a higher melting point compared to FA and its asymmetric dimers. Antioxidant and anti-proliferative assessments indicated that enzymatic oligomerization increased antioxidant and anti-proliferative properties of oxidation products for P2, P3 and P6 compared to FA. Finally, this enzymatic process in water could produce new molecules, having goo...
A new glyco-phenol was produced by the coupling between glucosamine (Glu) and ferulic acid (FA) u... more A new glyco-phenol was produced by the coupling between glucosamine (Glu) and ferulic acid (FA) using Myceliophthora thermophila laccase as biocatalyst in mild conditions (distilled water and 30°C) as an environmentally friendly process. Results indicated that the enzymatic reaction created a new derivative (FA-Glu), produced from coupling between Glu and FA by covalent bonds. By the high production of (FA-Glu) derivative and its stability, the optimal ratio of (FA: Glu) was of (1:1) at optimal time reaction of 6 h. Under these optimal conditions, almost 55% of -NH2 groups on Glu were bound with FA-oxidation products. The new derivative showed higher hydrophobic character than Glu due to the presence of FA in its structure. LC-MS analysis showed that (FA-Glu) derivative exhibited a molecular mass at MM 713 g/mol containing one Glu-molecule and three FA-molecules after decarboxylation. Furthermore, the new derivative presented good antioxidant and anti-proliferative activities in comparison with Glu and FA. These results suggest that the enzymatic conjugation between Glu and FA is a promising process to produce a new glyco-phenol having good functional properties for potential applications. This article is protected by copyright. All rights reserved.
Many studies underline the great benefits of yeast extracts (YE), used as supplements in animal f... more Many studies underline the great benefits of yeast extracts (YE), used as supplements in animal free culture media, on cell growth and recombinant protein production [1,2]. Nevertheless, their unknown composition and batch-to-batch variability of commercial YE remain constraints for industrial processes [3,4]. Consequently, the identification of bioactive YE molecules is challenging for process reliability. The main strategy to respond upon this problem is to fractionate the extract and then to characterize the fractions. So far, several fractionation processes such as ultrafiltration [5], gel filtration chromatography [6] or alcoholic precipitation [7] have been investigated. These studies suggested that the active components were mainly small molecules (< 1000 Da). But, the other physico-chemical properties of YE components have been poorly studied until now. In regards of this statement, it is proposed to get better knowledge of the YE molecules leading to an improvement of CH...
All in-text references underlined in blue are linked to publications on ResearchGate, letting you... more All in-text references underlined in blue are linked to publications on ResearchGate, letting you access and read them immediately.
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