Strategies based on aqueous two-phase systems for the separation of laccase from protease produced by Pleurotus ostreatus

Sánchez‐Trasviña, Calef; Enriquez-Ochoa, Daniela; Arellano-Gurrola, Clarisa Michelle; Tinoco‐Valencia, Raunel; Rito‐Palomares, Marco; Serrano‐Carreón, Leobardo; Mayolo‐Deloisa, Karla

doi:10.1016/j.fluid.2019.112281

Cited by 9 publications

(17 citation statements)

References 35 publications

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“…This value follows the gradual increase in the protein partition coefficient K P , which determines that the purification process occurs especially due to the migration of contaminating proteins to the polymeric phase (top phase). These behaviors were also observed in the studies by Sánchez-Trasviña et al 18 when Pleurotus ostreatus contaminating proteins tended to the polymeric phase by isolating the Lac in the bottom phase by increasing the concentration of PEG. A similar fact was described by Bertrand et al 33 who, when purifying the P. ostratus Lac in an LLE formed by the thermoseparable copolymer UCON and ammonium sulfate, observed that the Lac was predominantly concentrated in the bottom phase (salt-rich phase).…”

Section: Effect Of Polymer Concentrationsupporting

confidence: 76%

“…15 Few studies relate to the purification of lignin-modifying enzymes in LLE, which mostly fit a specificity of the LLE using ATPSs. 10,[16][17][18] However, it may be strategically advantageous to utilize phaseforming components that are not completely miscible in water. This approach results in the formation of liquid-liquid systems, which may be more suitable for the integrated stages of fermentation processes.…”

Section: Introductionmentioning

confidence: 99%

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Thermosensitive polymer‐assisted extraction and purification of fungal laccase from citrus pulp wash effluent

Santana,

Barros,

Canuto

et al. 2023

J Sci Food Agric

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BackgroundThis study explores the use of liquid‐liquid extraction (LLE) with thermosensitive polymers for producing laccase (Lac) from Pleurotus sajor‐caju. This process leverages liquid waste from the citrus industry, specifically pulp wash. The research delves into extractive fermentation and thermoseparation, both processes were facilitated by a polymer exhibiting a lower critical solution temperature (LCST) transition.ResultsKey factors considered include the choice of polymer, its concentration, pH, separation temperature, and the behavior of the polymer‐rich phase post‐extractive fermentation concerning LCST. Notably, under conditions of 45% by weight of Pluronic L‐61 and a pH of 5.0 at 25°C, the laccase was achieved an enhancement in the purification factor (PF) of 28.4‐fold, compared to the laccase obtained directly from the fermentation process on the 8th day. There was an 83.6% recovery of the laccase enzyme in the bottom phase of the system. Additionally, the unique properties of Pluronic L‐61, which can induce phase separation and also allow for thermoseparation, led to a secondary fraction (aqueous solution) of laccase with PF of 2.1 ± 0.1‐fold (at 32 ± 0.9 °C and 30 ± 0.3 min without stirring) from polymeric phase (top‐phase). The FTIR analysis validated the separation data, particularly highlighting the α‐helix content in the Amide I region (1600 ‐ 1700 cm‐1).ConclusionIn summary, the insights from this study pave the way for broader industrial applications of these techniques, underscoring benefits like streamlined process integration, heightened selectivity, and superior separation efficacy.This article is protected by copyright. All rights reserved.

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Section: Effect Of Polymer Concentrationsupporting

confidence: 76%

Section: Introductionmentioning

confidence: 99%

Thermosensitive polymer‐assisted extraction and purification of fungal laccase from citrus pulp wash effluent

Santana,

Barros,

Canuto

et al. 2023

J Sci Food Agric

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“…In PEG6000–citrate and PEG8000–citrate ATPS at V R 1.0, TLL had opposite effects on HA recovery depending on the MW of the polysaccharide. In general, as TLL increased, the free volume in the top phase was reduced (Sánchez-Trasviña et al 2019 ); thus, HA was forced to migrate towards the bottom phase as TLL increased. This was observed for sample HA3.5 but not for HA1.5.…”

Section: Discussionmentioning

confidence: 99%

“…ATPS, given their aqueous environment and mild conditions, have been largely employed for the recovery and separation of a broad range of biomacromolecules, including proteins, enzymes, antibodies, peptides, and genetic material (Asenjo and Andrews 2011 ; Azevedo et al 2009 ; Sánchez-Trasviña et al 2019 ), as well as low molecular weight compounds with biological activity (Enriquez-Ochoa et al 2020 ; Ghaffari et al 2019 ; Simental-Martínez et al 2014 ). Nevertheless, the application of ATPS for the separation of polysaccharides has been limited, mainly oriented as an extraction technique for polysaccharides from plant sources using ethanol-salt ATPS (Chen et al 2016 ; Cheng et al 2017 ; Wu et al 2017 ; Zhang et al 2018 ; Zhu et al 2020 ), whereas the implementation of ATPS in the downstream processing of polysaccharides from microbial fermentations is practically unexplored.…”

Section: Introductionmentioning

confidence: 99%

Primary recovery of hyaluronic acid produced in Streptococcus equi subsp. zooepidemicus using PEG–citrate aqueous two-phase systems

et al. 2021

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Given its biocompatibility, rheological, and physiological properties, hyaluronic acid (HA) has become a biomaterial of increasing interest with multiple applications in medicine and cosmetics. In recent decades, microbial fermentations have become an important source for the industrial production of HA. However, due to its final applications, microbial HA must undergo critical and long purification processes to ensure clinical and cosmetic grade purity. Aqueous two-phase systems (ATPS) have proven to be an efficient technique for the primary recovery of high-value biomolecules. Nevertheless, their implementation in HA downstream processing has been practically unexplored. In this work, polyethylene glycol (PEG)–citrate ATPS were used for the first time for the primary recovery of HA produced with an engineered strain of Streptococcus equi subsp. zooepidemicus. The effects of PEG molecular weight (MW), tie-line length (TLL), volume ratio (VR), and sample load on HA recovery and purity were studied with a clarified fermentation broth as feed material. HA was recovered in the salt-rich bottom phase, and its recovery increased when a PEG MW of 8000 g mol−1 was used. Lower VR values (0.38) favoured HA recovery, whereas purity was enhanced by a high VR (3.50). Meanwhile, sample load had a negative impact on both recovery and purity. The ATPS with the best performance was PEG 8000 g mol−1, TLL 43% (w/w), and VR 3.50, showing 79.4% HA recovery and 74.5% purity. This study demonstrated for the first time the potential of PEG–citrate ATPS as an effective primary recovery strategy for the downstream process of microbial HA.

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“…Laccase is a multicopper enzyme that can catalyze the oxidation of diverse compounds (e.g., phenols, polyphenols, amines, and various inorganic compounds) in the presence of mediators. [ 16 ] Laccase possesses four Cu 2+ ions per active monomer, and produces its oxidative effect through single‐electron oxidation and the simultaneous reduction of molecular oxygen (O 2 ) to yield two molecules of water. [ 17 ] Cations such as Cu 2+ and their complexes reportedly can catalyze luminol oxidation, and laccase‐mediated luminol oxidation can result in light emission.…”

Section: Introductionmentioning

confidence: 99%

Laccase–luminol chemiluminescence system: an investigation of substrate inhibition

et al. 2023

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Chemiluminescence (CL) reactions are widely used for the detection and quantification of many types of analytes. Laccase has previously been proposed in CL reactions; however, its light emission behaviour has not been characterized. This study was conducted to characterize the laccase–luminol system, determine its kinetic parameters, and analyze the effects of protein and OHˉ concentration on the CL signal. Laccase from Coriolopsis gallica was combined with different concentrations of luminol (125 nM to 4 mM), and the enzyme kinetics were evaluated using diverse kinetic models. The laccase–luminol system was able to produce CL without an intermediate molecule, but it exhibited substrate‐inhibition behaviour. A two‐site random model was used and suggested that when the first luminol molecule was bound to the active site, laccase affinity for the second luminol molecule was increased. This inhibition effect could be avoided using a low luminol concentration. At 5 μM luminol concentration, 1 mg/ml (0.13 U) laccase is needed to achieve nearly 90% of the maximum CL signal, suggesting that the available luminol could not bind to all active sites. Furthermore, the concentration of NaOH negatively affected the CL signal. The laccase–luminol system represents an alternative to existing CL systems, with potential uses in molecular detection and quantification.

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Strategies based on aqueous two-phase systems for the separation of laccase from protease produced by Pleurotus ostreatus

Cited by 9 publications

References 35 publications

Thermosensitive polymer‐assisted extraction and purification of fungal laccase from citrus pulp wash effluent

Thermosensitive polymer‐assisted extraction and purification of fungal laccase from citrus pulp wash effluent

Primary recovery of hyaluronic acid produced in Streptococcus equi subsp. zooepidemicus using PEG–citrate aqueous two-phase systems

Laccase–luminol chemiluminescence system: an investigation of substrate inhibition

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