A robotic platform to screen aqueous two-phase systems for overcoming inhibition in enzymatic reactions

Bussamra, Bianca Consorti; Gomes, Joana Castro; Freitas, Sindélia; Mussatto, Solange I.; Costa, Aline Carvalho da; Wielen, Luuk A.M. van der; Ottens, Marcel

doi:10.1016/j.biortech.2019.01.136

Cited by 10 publications

(10 citation statements)

References 42 publications

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“…The selection of ATPS composition (salt type and polymer molecular weight) and concentration (tie line length–TLL) were based on data previously published by Bussamra et al ( 2019 ). The ATPS, respective TLL, and experiment to which they were applied and motivation are presented in Table 2 .…”

Section: Methodsmentioning

confidence: 99%

“…Sugar and protein present in top and bottom phases were measured according to methodology described by Bussamra et al ( 2019 ). Sugar quantification followed the Megazyme Glucose Reagent assay (Megazyme, Wicklow, Ireland), and no adaptation was needed to ATPS samples.…”

Section: Methodsmentioning

confidence: 99%

“…Hydrolysis samples for glucose measurement were boiled for 5 min and 99°C after collected (Eppendorf Thermomixer® C.), in order to deactivate the enzymes. The protocol used for protein quantification, based on Bradford ( 1976 ) method and performed using the Coomassie Protein Assay Reagent (Thermo Scientific, USA), required adaptation for ATPS samples, explained in detail in the cited reference (Bussamra et al, 2019 ). The solute concentrations in g/L were defined per volume of the respective phase being measured.…”

Section: Methodsmentioning

confidence: 99%

“…Systems were formed by addition of the required amount of PEG and salt stock solutions to an aqueous solution of 5 mL total volume. Sugar stock solution (800 g/L, prepared in Milli-Q water) was added to a (Bussamra et al, 2019).…”

Section: Atps Formation (Equilibrium and Separation)mentioning

confidence: 99%

“…Another limitation of ATPS application relies on the analytical techniques to quantify proteins (González-González 200, 116.3, 97.4, 66.3, 55.4, 36.5, 31, 21.5, 14.4, 6, 3.5, and 2.5. et al, 2011;Silvério et al, 2012;Glyk et al, 2015). Some strategies have already been suggested in literature using conventional methods of quantification (Bussamra et al, 2019).…”

Section: Atps and Enzymatic Hydrolysis Of Sugarcane Bagassementioning

confidence: 99%

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Enzymatic Hydrolysis of Sugarcane Bagasse in Aqueous Two-Phase Systems (ATPS): Exploration and Conceptual Process Design

et al. 2020

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The enzymatic conversion of lignocellulosic material to sugars can provide a carbon source for the production of energy (fuels) and a wide range of renewable products. However, the efficiency of this conversion is impaired due to product (sugar) inhibition. Even though several studies investigate how to overcome this challenge, concepts on the process to conduct the hydrolysis are still scarce in literature. Aqueous two-phase systems (ATPS) can be applied to design an extractive reaction due to their capacity to partition solutes to different phases in such a system. This work presents strategies on how to conduct extractive enzymatic hydrolysis in ATPS and how to explore the experimental results in order to design a feasible process. While only a limited number of ATPS was explored, the methods and strategies described could easily be applied to any further ATPS to be explored. We studied two promising ATPS as a subset of a previously high throughput screened large set of ATPS, providing two configurations of processes having the reaction in either the top phase or in the bottom phase. Enzymatic hydrolysis in these ATPS was performed to evaluate the partitioning of the substrate and the influence of solute partitioning on conversion. Because ATPS are able to partition inhibitors (sugar) between the phases, the conversion rate can be maintained. However, phase forming components should be selected to preserve the enzymatic activity. The experimental results presented here contribute to a feasible ATPS-based conceptual process design for the enzymatic conversion of lignocellulosic material.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Atps Formation (Equilibrium and Separation)mentioning

confidence: 99%

Section: Atps and Enzymatic Hydrolysis Of Sugarcane Bagassementioning

confidence: 99%

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Enzymatic Hydrolysis of Sugarcane Bagasse in Aqueous Two-Phase Systems (ATPS): Exploration and Conceptual Process Design

et al. 2020

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Anything but Conventional Chromatography Approaches in Bioseparation

Roque

Pina

Azevedo

et al. 2020

Biotechnology Journal

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While packed bed chromatography, known as conventional chromatography, has been serving the biopharmaceutical industry for decades as the bioseparation method of choice, alternative approaches are likely to take an increasing leading role in the next few years. The high number of new biological drugs under development, and the need to make biopharmaceuticals widely accessible, has been driving the academia and industry in the quest of anything but conventional chromatography approaches. In this perspective paper, these alternative approaches are discussed in view of current and future challenges in the downstream processing field.

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Automation and miniaturization: enabling tools for fast, high‐throughput process development in integrated continuous biomanufacturing

Silva

Eppink

Ottens

2021

J of Chemical Tech & Biotech

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Process development in the biotech industry leads to investments around hundred of millions of dollars. It is important to mitigate costs without neglecting the quality of process development. Biopharmaceutical process development is important for companies to develop new processes and be first to market, improve a pre‐established process, or start manufacturing a product available by patent expiry (biosimilars). Laboratory automation enables methodical and standardized process development. Miniaturization and parallelization empower laboratories to screen several experimental conditions and define operating windows for purification processes, improving process robustness. Together, they allow for fast and accurate process development in a fraction of the time and cost of nonminiaturized/nonparallel process development approaches. The most widely used High‐Throughput Screening technique is a liquid‐handling station and microfluidics is taking its first steps in process development. Both are attractive scale‐down tools for the characterization of bioprocesses and allow thousands of experiments to be performed per day. High‐Throughput Process Development (HTPD) has helped to achieve major breakthroughs in process optimization, both for upstream and downstream processing. Continuous processing is the next step in process development which leads to cost reduction, higher productivity and better quality control; the integration of upstream and downstream processes is seen as a major challenge. In this review, we will focus on the state‐of‐the‐art of miniaturized techniques for process development in the biotechnology industry, and how automation and miniaturization drive process development. A comparison between liquid‐handling stations and microfluidics is made and an indication is given of which tools are still lacking for HTPD in the context of Integrated Continuous Biomanufacturing. © 2021 The Authors. Journal of Chemical Technology and Biotechnology published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry (SCI).

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A robotic platform to screen aqueous two-phase systems for overcoming inhibition in enzymatic reactions

Cited by 10 publications

References 42 publications

Enzymatic Hydrolysis of Sugarcane Bagasse in Aqueous Two-Phase Systems (ATPS): Exploration and Conceptual Process Design

Enzymatic Hydrolysis of Sugarcane Bagasse in Aqueous Two-Phase Systems (ATPS): Exploration and Conceptual Process Design

Anything but Conventional Chromatography Approaches in Bioseparation

Automation and miniaturization: enabling tools for fast, high‐throughput process development in integrated continuous biomanufacturing

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