Journal of Chemical Technology and Biotechnology, 2019
Traditionally, aqueous two‐phase systems (ATPS) have been used as a liquid–liquid extraction tech... more Traditionally, aqueous two‐phase systems (ATPS) have been used as a liquid–liquid extraction technique for the primary recovery and purification of biological samples. The enormous potential of their usage comes with great economical and technical advantages mainly due to the mild physicochemical environment. Nowadays, the use of ATPS as a bioengineering technique is approaching an era where new possibilities are being explored to maximize their use and implementation in the development of novel practical applications and tools. In this context, ‘intelligent’ polymers are being used as phase forming chemicals in ATPS in route to process integration. Extractive fermentation in ATPS is being re‐evaluated with the aim of effectively growing microorganisms while recovering their fermentation products in different phases. ATPS are also being used as a tool for refolding of proteins. There are also several innovative efforts being made towards implementing this bioengineering tool as a continuous process moving away from traditional batch operations. In general, the possibilities of implementing ATPS in different bioprocessing aspects are growing considerably and gaining importance. This review aims to present the novel trends in the use and development of ATPS strategies as complete bioengineering tools and to provide a full perspective of their possibilities in the near future.
Uricase is the enzyme responsible for the breakdown of uric acid, the key molecule leading to gou... more Uricase is the enzyme responsible for the breakdown of uric acid, the key molecule leading to gout in humans, into allantoin, but it is absent in humans. It has been produced as a PEGylated pharmaceutical where the purification is performed through three sequential chromatographic columns. More recently an aqueous two-phase system (ATPS) was reported that could recover Uricase with high yield and purity. Although the use of ATPS can decrease cost and time, it also generates a large amount of waste. The ability, therefore, to recycle key components of ATPS is of interest. Economic modelling is a powerful tool that allows the bioprocess engineer to compare possible outcomes and find areas where further research or optimization might be required without recourse to extensive experiments and time. This research provides an economic analysis using the commercial software BioSolve of the strategies for Uricase production: chromatographic and ATPS, and includes a third bioprocess that utilises material recycling. The key parameters that affect the process the most were located via a sensitivity analysis and evaluated with a Monte Carlo analysis. Results show that ATPS is far less expensive than chromatography, but that there is an area where the cost of production of both bioprocesses overlap. Furthermore, recycling doesn't impact the cost of production. This study serves to provide a framework for the economic analysis of Uricase production using alternative techniques. This article is protected by copyright. All rights reserved.
The potential recovery of high-value products from brewery yeast waste confers value to this indu... more The potential recovery of high-value products from brewery yeast waste confers value to this industrial residue. Aqueous two-phase systems (ATPS) have demonstrated to be an attractive alternative for the primary recovery of biological products and are therefore suitable for the recovery of invertase from this residue. Sixteen different polyethylene glycol (PEG)–potassium phosphate ATPS were tested to evaluate the effects of PEG molecular weight (MW) and tie-line length (TLL) upon the partition behavior of invertase. Concentrations of crude extract from brewery yeast waste were then varied in the systems that presented the best behaviors to intensify the potential recovery of the enzyme. Results show that the use of a PEG MW 400 g mol −1 system with a TLL of 45.0% (w/w) resulted in an invertase bottom phase recovery with a purification factor of 29.5 and a recovery yield of up to 66.2% after scaling the system to a total weight of 15.0 g. This represents 15.1 mg of invertase per mL of processed bottom phase. With these results, a single-stage ATPS process for the recovery of invertase is proposed.
Journal of chromatography. B, Biomedical applications, Jan 17, 1996
A practical study is presented of the influence of cell debris and polymer recycling upon the ope... more A practical study is presented of the influence of cell debris and polymer recycling upon the operation of two-stage aqueous two-phase systems (ATPS) for the recovery of yeast bulk protein, pyruvate kinase and fumarase. Brewers' yeast was disrupted using one of two types of high-pressure homogenisers or a bead mill. The different cell debris suspensions were partitioned in a single PEG-phosphate ATPS extraction and the efficiency of solid-liquid separation was examined. A continuously operated two-stage ATPS process, using spray columns, is presented and practical problems of polymer recycling are discussed. Conclusions are drawn concerning the generic implementation and operational stability of ATPS in practical protein recoveries.
Journal of Chemical Technology and Biotechnology, 2019
Traditionally, aqueous two‐phase systems (ATPS) have been used as a liquid–liquid extraction tech... more Traditionally, aqueous two‐phase systems (ATPS) have been used as a liquid–liquid extraction technique for the primary recovery and purification of biological samples. The enormous potential of their usage comes with great economical and technical advantages mainly due to the mild physicochemical environment. Nowadays, the use of ATPS as a bioengineering technique is approaching an era where new possibilities are being explored to maximize their use and implementation in the development of novel practical applications and tools. In this context, ‘intelligent’ polymers are being used as phase forming chemicals in ATPS in route to process integration. Extractive fermentation in ATPS is being re‐evaluated with the aim of effectively growing microorganisms while recovering their fermentation products in different phases. ATPS are also being used as a tool for refolding of proteins. There are also several innovative efforts being made towards implementing this bioengineering tool as a continuous process moving away from traditional batch operations. In general, the possibilities of implementing ATPS in different bioprocessing aspects are growing considerably and gaining importance. This review aims to present the novel trends in the use and development of ATPS strategies as complete bioengineering tools and to provide a full perspective of their possibilities in the near future.
Uricase is the enzyme responsible for the breakdown of uric acid, the key molecule leading to gou... more Uricase is the enzyme responsible for the breakdown of uric acid, the key molecule leading to gout in humans, into allantoin, but it is absent in humans. It has been produced as a PEGylated pharmaceutical where the purification is performed through three sequential chromatographic columns. More recently an aqueous two-phase system (ATPS) was reported that could recover Uricase with high yield and purity. Although the use of ATPS can decrease cost and time, it also generates a large amount of waste. The ability, therefore, to recycle key components of ATPS is of interest. Economic modelling is a powerful tool that allows the bioprocess engineer to compare possible outcomes and find areas where further research or optimization might be required without recourse to extensive experiments and time. This research provides an economic analysis using the commercial software BioSolve of the strategies for Uricase production: chromatographic and ATPS, and includes a third bioprocess that utilises material recycling. The key parameters that affect the process the most were located via a sensitivity analysis and evaluated with a Monte Carlo analysis. Results show that ATPS is far less expensive than chromatography, but that there is an area where the cost of production of both bioprocesses overlap. Furthermore, recycling doesn't impact the cost of production. This study serves to provide a framework for the economic analysis of Uricase production using alternative techniques. This article is protected by copyright. All rights reserved.
The potential recovery of high-value products from brewery yeast waste confers value to this indu... more The potential recovery of high-value products from brewery yeast waste confers value to this industrial residue. Aqueous two-phase systems (ATPS) have demonstrated to be an attractive alternative for the primary recovery of biological products and are therefore suitable for the recovery of invertase from this residue. Sixteen different polyethylene glycol (PEG)–potassium phosphate ATPS were tested to evaluate the effects of PEG molecular weight (MW) and tie-line length (TLL) upon the partition behavior of invertase. Concentrations of crude extract from brewery yeast waste were then varied in the systems that presented the best behaviors to intensify the potential recovery of the enzyme. Results show that the use of a PEG MW 400 g mol −1 system with a TLL of 45.0% (w/w) resulted in an invertase bottom phase recovery with a purification factor of 29.5 and a recovery yield of up to 66.2% after scaling the system to a total weight of 15.0 g. This represents 15.1 mg of invertase per mL of processed bottom phase. With these results, a single-stage ATPS process for the recovery of invertase is proposed.
Journal of chromatography. B, Biomedical applications, Jan 17, 1996
A practical study is presented of the influence of cell debris and polymer recycling upon the ope... more A practical study is presented of the influence of cell debris and polymer recycling upon the operation of two-stage aqueous two-phase systems (ATPS) for the recovery of yeast bulk protein, pyruvate kinase and fumarase. Brewers' yeast was disrupted using one of two types of high-pressure homogenisers or a bead mill. The different cell debris suspensions were partitioned in a single PEG-phosphate ATPS extraction and the efficiency of solid-liquid separation was examined. A continuously operated two-stage ATPS process, using spray columns, is presented and practical problems of polymer recycling are discussed. Conclusions are drawn concerning the generic implementation and operational stability of ATPS in practical protein recoveries.
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