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MICHAEL HOARE - One of the best experts on this subject based on the ideXlab platform.

  • An ultra scale‐down characterization of low shear stress Primary Recovery stages to enhance selectivity of fusion protein Recovery from its molecular variants
    Biotechnology and Bioengineering, 2013
    Co-Authors: Simyee Kong, Shaun Mcnulty, Claire Entwisle, Ann Mcilgorm, Kate A. Dalton, MICHAEL HOARE
    Abstract:

    Fusion proteins offer the prospect of new therapeutic products with multiple functions. The Primary Recovery is investigated of a fusion protein consisting of modified E2 protein from hepatitis C virus fused to human IgG1 Fc and expressed in a Chinese hamster ovary (CHO) cell line. Fusion protein products inevitably pose increased challenge in preparation and purification. Of particular concerns are: (i) the impact of shear stress on product integrity and (ii) the presence of product-related contaminants which could prove challenging to remove during the high resolution purification steps. This paper addresses the use of microwell-based ultra scale-down (USD) methods to develop a bioprocess strategy focused on the integration of cell culture and cell removal operations and where the focus is on the use of operations which impart low shear stress levels even when applied at eventual manufacturing scale. An USD shear device was used to demonstrate that cells exposed to high process stresses such as those that occur in the feed zone of a continuous non-hermetic centrifuge resulted in the reduction of the fusion protein and also the release of glycosylated intracellular variants. In addition, extended cell culture resulted in release of such variants. USD mimics of low shear stress, hydrohermetic feed zone centrifugation and of depth filtration were used to demonstrate little to no release during Recovery of these variants with both results verified at pilot scale. Furthermore, the USD studies were used to predict removal of contaminants such as lipids, nucleic acids, and cell debris with, for example, depth filtration delivering greater removal than for centrifugation but a small (∼10%) decrease in yield of the fusion protein. These USD observations of product Recovery and carryover of contaminants were also confirmed at pilot scale as was also the capacity or throughput achievable for continuous centrifugation or for depth filtration. The advantages are discussed of operating a lower yield cell culture and a low shear stress Recovery process in return for a considerably less challenging purification demand. Biotechnol. Bioeng. 2013; 110: 1973–1983. © 2013 Wiley Periodicals, Inc.

  • An ultra scale-down characterization of low shear stress Primary Recovery stages to enhance selectivity of fusion protein Recovery from its molecular variants.
    Biotechnol Bioeng, 2013
    Co-Authors: MICHAEL HOARE
    Abstract:

    Fusion proteins offer the prospect of new therapeutic products with multiple functions. The Primary Recovery is investigated of a fusion protein consisting of modified E2 protein from hepatitis C virus fused to human IgG1 Fc and expressed in a Chinese hamster ovary (CHO) cell line. Fusion protein products inevitably pose increased challenge in preparation and purification. Of particular concerns are: (i) the impact of shear stress on product integrity and (ii) the presence of product-related contaminants which could prove challenging to remove during the high resolution purification steps. This paper addresses the use of microwell-based ultra scale-down (USD) methods to develop a bioprocess strategy focused on the integration of cell culture and cell removal operations and where the focus is on the use of operations which impart low shear stress levels even when applied at eventual manufacturing scale. An USD shear device was used to demonstrate that cells exposed to high process stresses such as those that occur in the feed zone of a continuous non-hermetic centrifuge resulted in the reduction of the fusion protein and also the release of glycosylated intracellular variants. In addition, extended cell culture resulted in release of such variants. USD mimics of low shear stress, hydrohermetic feed zone centrifugation and of depth filtration were used to demonstrate little to no release during Recovery of these variants with both results verified at pilot scale. Furthermore, the USD studies were used to predict removal of contaminants such as lipids, nucleic acids, and cell debris with, for example, depth filtration delivering greater removal than for centrifugation but a small (~10%) decrease in yield of the fusion protein. These USD observations of product Recovery and carryover of contaminants were also confirmed at pilot scale as was also the capacity or throughput achievable for continuous centrifugation or for depth filtration. The advantages are discussed of operating a lower yield cell culture and a low shear stress Recovery process in return for a considerably less challenging purification demand.

Jorge Benavides - One of the best experts on this subject based on the ideXlab platform.

  • Primary Recovery of bioactive compounds from stressed carrot tissue using aqueous two-phase systems strategies
    Journal of Chemical Technology & Biotechnology, 2014
    Co-Authors: Juan Carlos Sánchez-rangel, Daniel A. Jacobo-velázquez, Luis Cisneros-zevallos, Jorge Benavides
    Abstract:

    BACKGROUND The application of controlled postharvest abiotic stresses in carrots induces the production of phenolic compounds, mainly chlorogenic acids, which present diverse biological functions. Aqueous two-phase systems (ATPS) is a liquid–liquid extraction technique that has demonstrated its potential for the fractionation and Recovery of bioactive compounds. ATPS have the advantages of using environmentally friendly constituents, ease of scale-up and process integration capability. This experimental work addresses for the first time the Recovery of bioactive compounds (mainly hydroxycinnamic acid derivatives) from abiotically stressed carrot tissue using ATPS strategies, including in situ extraction, in alcohol–salt and ionic liquid–salt systems. RESULTS The partitioning of chlorogenic acid was favored in the ethanol–potassium phosphate system achieving a Recovery yield of 81.9%. Hydroxycinnamic acid derivatives fractioning was favored in 1-ethyl-3-methylimidazolium acetate (EMIM-Ac)–potassium phosphate ATPS, where Recovery yields up to 85.4% were achieved. The in situ extraction strategy not only demonstrated potential for merging extraction and Recovery in the same system but also, depending on ATPS used, allowed differential phenolic profiles to be obtained. CONCLUSION ATPS strategies (either solid–liquid extraction followed by ATPS or in situ extraction) were shown to have great potential for the Recovery of bioactives from stressed carrot tissue. © 2014 Society of Chemical Industry

  • rotavirus like particles Primary Recovery from insect cells in aqueous two phase systems
    Journal of Chromatography B, 2006
    Co-Authors: Jorge Benavides, Jimmy A Mena, Mayra Cisnerosruiz, Octavio T Ramirez, Laura A Palomares, Marco Ritopalomares
    Abstract:

    Abstract Virus-like particles have a wide range of applications, including vaccination, gene therapy, and even as nanomaterials. Their successful utilization depends on the availability of selective and scalable methods of product Recovery and purification that integrate effectively with upstream operations. In this work, a strategy based on aqueous two phase system (ATPS) was developed for the Recovery of double-layered rotavirus-like particles (dlRLP) produced by the insect cell-baculovirus expression system. Polyethylene glycol (PEG) molecular mass, PEG and salt concentrations, and volume ratio (Vr, volume of top phase/volume of bottom phase) were evaluated in order to determine the conditions where dlRLP and contaminants concentrated to opposite phases. Two-stage ATPS consisting of PEG 400-phosphate with a Vr of 13.0 and a tie-line length (TLL) of 35% (w/w) at pH 7.0 provided the best conditions for processing highly concentrated crude extract from disrupted cells (dlRLP concentration of 5 μg/mL). In such conditions intracellular dlRLP accumulated in the top phase (Recovery of 90%), whereas cell debris remained in the interface. Furthermore, dlRLP from culture supernatants accumulated preferentially in the interface (Recovery of 82%) using ATPS with a Vr of 1.0, pH of 7.0, PEG 3350 (10.1%, w/w) and phosphate (10.9%, w/w). The purity of dlRLP from culture supernatant increased up to 55 times after ATPS. The use of ATPS resulted in a Recovery process that produced dlRLP with a purity between 6 and 11% and an overall product yield of 85% (w/w), considering purification from intracellular and extracellular dlRLP. Overall, the strategy proposed in this study is simpler than traditional methods for recovering dlRLP, and represents a scalable and economically viable alternative for production processes of vaccines against rotavirus infection with significant scope for generic commercial application.

  • Potential Aqueous Two-Phase Processes for the Primary Recovery of Colored Protein from Microbial Origin
    Engineering in Life Sciences, 2005
    Co-Authors: Jorge Benavides, Marco Rito-palomares
    Abstract:

    The Primary Recovery of c-phycocyanin and b-phycoerythrin from Spirulina maxima and Porphyridium cruentum, respectively, using an established extraction strategy was selected as a practical model system to study the generic application of polyethylene glycol (PEG)-phosphate aqueous two-phase systems (ATPS). The generic practical implementation of ATPS extraction was evaluated for the Recovery of colored proteins from microbial origin. A comparison of the influence of system parameters, such as PEG molecular mass, concentration of PEG as well as salt, system pH and volume ratio, on the partition behavior of c-phycocyanin and b-phycoerythrin was carried out to determine under which conditions target colored protein and contaminants concentrate to opposite phases. One-stage processes are proposed for the Primary Recovery of the colored proteins. PEG1450-phosphate ATPS extraction (volume ratio (VR) equal to 0.3, tie-line length (TLL) of 34 % w/w and system pH 7.0) for the Recovery of c-phycocyanin from Spirulina maxima resulted in a Primary Recovery process that produced a protein purity of 2.1 ± 0.2 (defined as the relationship of 620 nm to 280 nm absorbance) and a product yield of 98 % [w/w]. PEG1000-phosphate ATPS extraction (i.e., VR = 1.0, PEG 1000, TLL 50 % w/w and system pH 7.0) was preferred for the Recovery of b-phycoerythrin from Porphyridium cruentum, which resulted in a protein purity of 2.8 ± 0.2 (defined as the relationship of 545 nm to 280 nm absorbance) and a product yield of 82 % [w/w]. The purity of c-phycocyanin and b-phycoerythrin from the crude extract increased 3- and 4-fold, respectively, after ATPS. The results reported herein demonstrated the benefits of the practical generic application of ATPS for the Primary Recovery of colored proteins from microbial origin as a first step for the development of purification processes.

  • Bioprocess intensification: a potential aqueous two-phase process for the Primary Recovery of B-phycoerythrin from Porphyridium cruentum.
    Journal of Chromatography B, 2004
    Co-Authors: Jorge Benavides, Marco Rito-palomares
    Abstract:

    Abstract A process for the Primary Recovery of B-phycoerythrin from Porphyridium cruentum exploiting aqueous two-phase systems (ATPS) was developed in order to reduce the number of unit operations and benefit from an increased yield of the protein product. The evaluation of system parameters such as poly(ethylene glycol) (PEG) molecular mass, concentration of PEG as well as salt, system pH and volume ratio was carried out to determine under which conditions the B-phycoerythrin and contaminants concentrate to opposite phases. PEG 1450-phosphate ATPS proved to be suitable for the Recovery of B-phycoerythrin because the target protein concentrated to the top phase whilst the protein contaminants and cell debris concentrated in the bottom phase. An extraction ATPS stage comprising volume ratio (Vr) equal to 1.0, PEG 1450 24.9% (w/w), phosphate 12.6% (w/w) and system pH of 8.0 allowed B-phycoerythrin Recovery with a purity of 2.9 (estimated as the relation of the 545–280 nm absorbances). The use of ATPS resulted in a Primary Recovery process that produced a protein purity of 2.9±0.2 and an overall product yield of 77.0% (w/w). The results reported demonstrated the practical implementation of ATPS for the design of a Primary Recovery process as a first step for the commercial purification of B-phycoerythrin produced by P. cruentum.

Arbakariya B. Ariff - One of the best experts on this subject based on the ideXlab platform.

  • Aqueous two-phase flotation for Primary Recovery of bacteriocin-like inhibitory substance (BLIS) from Pediococcus acidilactici Kp10.
    Journal of Chromatography B, 2016
    Co-Authors: Nurul Lyana Md Sidek, Sahar Abbasiliasi, Fadzlie Wong Faizal Wong, Shuhaimi Mustafa, Arbakariya B. Ariff
    Abstract:

    Abstract An aqueous two-phase flotation (ATPF) system based on polyethylene glycol (PEG) and sodium citrate (NaNO 3 C 6 H 5 O 7 ·2H 2 O) was considered for Primary Recovery of bacteriocin-like inhibitory substance (BLIS) from Pediococcus acidilactici Kp10. The effects of ATPF parameters namely phase composition, tie-line length (TLL), volume ratio between the two phases (V R ), amount of crude load (C L ), pH, nitrogen gas flow rate (F R ) and flotation time (F T ) on the performance of Recovery were evaluated. BLIS was mainly concentrated into the upper PEG-rich phase in all systems tested so far. The optimum conditions for BLIS purification, which composed of PEG 8000/sodium citrate, were: TLL of 42.6, V R of 0.4, C L of 22% (w/w), pH 7, average F T of 30 min and F R of 20 mL/min. BLIS was partially purified up to 5.9-fold with a separation efficiency of 99% under this optimal conditions. A maximum yield of BLIS activity of about 70.3% was recovered in the PEG phase. The BLIS from the top phase was successfully recovered with a single band in SDS-gel with molecular weight of about 10–15 kDa. ATPF was found to be an effective technique for the Recovery of BLIS from the fermentation broth of P. acidilactici Kp10.

  • Primary Recovery of miraculin from miracle fruit, Synsepalum dulcificum by AOT reverse micellar system
    Lwt - Food Science and Technology, 2015
    Co-Authors: Zuxing He, Sahar Abbasiliasi, Murni Halim, Arbakariya B. Ariff
    Abstract:

    Abstract Miracle fruit, Synsepalum dulcificum , contains a glycoprotein known as miraculin. After consuming this glycoprotein, sour foods taste sweet and the effect lasts for up to 4 h. With increasing demand for natural and “low-calorie” sweeteners, the use of miraculin as an additive is increasing enormously in the food, medicine and cosmetic industries. In this study, we used reverse micelles formed from a sodium di (2-ethylhexyl) sulfosuccinate (AOT)/isooctane system to purify miraculin from S. dulcificum. We studied factors affecting purification performance, such as surfactant (AOT) concentration and the pH of the crude during forward extraction. During backward extraction, we examined the effects of NaCl concentration, the pH of the aqueous phase and addition of isopropanol. We found that 0.1 mol/L AOT/isooctane solution mixed with crude extract at pH 8 during the forward extraction stage and 0.5 mol/L NaCl solution at pH 11 during the backward stripping stage were optimal purification conditions, from which 22% miraculin was recovered with a purity of 94.8%.

  • Primary Recovery of thermostable lipase 42 derived from recombinant Escherichia coli BL21 in aqueous two-phase flotation
    Separation and Purification Technology, 2014
    Co-Authors: Sahar Abbasiliasi, Mohd Shamzi Mohamed, Mohammad Rizal Kapri, Saeid Kadkhodaei, Raja Noor Zaliha Raja Abd Rahman, Arbakariya B. Ariff
    Abstract:

    Abstract An aqueous two-phase flotation (ATPF) composed of polyethylene glycol (PEG) and sodium citrate was constructed for direct purification of thermostable lipase 42 from recombinant Escherichia coli BL21(DE3) pLysS. The influences of varying phase forming composition, tie-line length (TLL), volume ratio ( V R ), crude loading ( C L ), system pH, gas nitrogen flow rate ( F R ) and flotation time ( F t ) upon ATPF partitioning performance were investigated. The optimum purification condition of lipase was achieved at PEG 8000/sodium citrate ATPF comprising TLL of 25.4, V R of 0.3, C L of 20% (w/w) at pH 7 with average F t of 10 min and F R at 20 mL/min. Lipase was successfully purified using ATPF up to 4.05-fold with a separation efficiency of 99%. Result of this study also demonstrated that Recovery yield of lipase in PEG phase of up to 96% was achieved. These results reveal the potential of ATPF partitioning in Recovery and purification of recombinant lipase.

  • Primary Recovery of a bacteriocin like inhibitory substance derived from pediococcus acidilactici kp10 by an aqueous two phase system
    Food Chemistry, 2014
    Co-Authors: Sahar Abbasiliasi, Tau Chuan Ling, Shuhaimi Mustafa, Saeid Kadkhodaei, Hui Suan Ng, Tengku Azmi Tengku Ibrahim, Faezeh Vakhshiteh, Zahra Ajdari, Raha Abdul Rahim, Arbakariya B. Ariff
    Abstract:

    A polymer–salt aqueous two-phase system (ATPS) consisting of polyethylene–glycol (PEG) with sodium citrate was developed for direct Recovery of a bacteriocin-like inhibitory substance (BLIS) from a culture of Pediococcus acidilactici Kp10. The influences of phase composition, tie-line length (TLL), volume ratio (VR), crude sample loading, pH and sodium chloride (NaCl) on the partition behaviour of BLIS was investigated. Under optimum conditions of ATPS, the purification of BLIS was achieved at 26.5% PEG (8000)/11% sodium citrate with a TLL of 46.38% (w/w), VR of 1.8, and 1.8% crude load at pH 7 without the presence of NaCl. BLIS from P. acidilactici Kp10 was successfully purified by the ATPS up to 8.43-fold with a yield of 81.18%. Given that the operation of ATPS is simple, environmentally friendly and cost-effective, as it requires only salts and PEG, it may have potential for industrial applications in the Recovery of BLIS from fermentation broth.

  • Primary Recovery of lipase derived from Burkholderia sp. ST8 with aqueous micellar two-phase system
    Process Biochemistry, 2011
    Co-Authors: Arbakariya B. Ariff, Shaliza Ibrahim, Tau Chuan Ling
    Abstract:

    The partitioning and Recovery of lipase derived from Burkholderia sp. ST8 strain was explored using temperature-induced aqueous micellar two-phase system (AMTPS) composed of single nonionic surfactant. Nonionic surfactant Triton X-114 and Pluronic series (triblock copolymer) were evaluated in terms of their clouding phenomenon (cloud-point temperature) and the performance of the lipase partitioning in these AMTPSs. Pluronic L81 showed the most optimum partition efficiency for the Recovery of lipase to the micellar phase of the AMTPS. Based on the AMTPS which consisted of 24% (w/w) Pluronic L81 and 0.5% (w/w) potassium chloride (KCl), the selectivity of lipase partitioned to bottom phase has been enhanced to 0.035 and the lipase was purified 7.2 fold. Furthermore, the lipase from the micellar phase was consecutively extracted to a new aqueous solution, with an aim of removing the surfactant from the purified lipase. It was attained by replacing the aqueous top phase from the Primary Recovery of AMTPS with a new potassium thiocyanate (KSCN) solution. The lipase was then recovered in the newly formed bottom aqueous phase which culminated in the yield of 89% and partition coefficients of 0.34 and 4.50 for lipase and surfactant, respectively. AMTPS offers a convenient and efficient method for the Primary Recovery of lipase with low cost, large loading capacity and the potential of linear scale up.

Nigel J. Titchener-hooker - One of the best experts on this subject based on the ideXlab platform.

  • Integrated economic and experimental framework for screening of Primary Recovery technologies for high cell density CHO cultures.
    Biotechnology Journal, 2016
    Co-Authors: Daria Popova, Adam Stonier, David Pain, Nigel J. Titchener-hooker, Suzanne S. Farid
    Abstract:

    Increases in mammalian cell culture titres and densities have placed significant demands on Primary Recovery operation performance. This article presents a methodology which aims to screen rapidly and evaluate Primary Recovery technologies for their scope for technically feasible and cost-effective operation in the context of high cell density mammalian cell cultures. It was applied to assess the performance of current (centrifugation and depth filtration options) and alternative (tangential flow filtration (TFF)) Primary Recovery strategies. Cell culture test materials (CCTM) were generated to simulate the most demanding cell culture conditions selected as a screening challenge for the technologies. The performance of these technology options was assessed using lab scale and ultra scale-down (USD) mimics requiring 25-110mL volumes for centrifugation and depth filtration and TFF screening experiments respectively. A centrifugation and depth filtration combination as well as both of the alternative technologies met the performance selection criteria. A detailed process economics evaluation was carried out at three scales of manufacturing (2,000L, 10,000L, 20,000L), where alternative Primary Recovery options were shown to potentially provide a more cost-effective Primary Recovery process in the future. This assessment process and the study results can aid technology selection to identify the most effective option for a specific scenario.

  • Integration of host strain bioengineering and bioprocess development using ultra‐scale down studies to select the optimum combination: An antibody fragment Primary Recovery case study
    Biotechnology and Bioengineering, 2014
    Co-Authors: Jean P. Aucamp, Richard Davies, Damien Hallet, Amanda Weiss, Nigel J. Titchener-hooker
    Abstract:

    An ultra scale-down Primary Recovery sequence was established for a platform E. coli Fab production process. It was used to evaluate the process robustness of various bioengineered strains. Centrifugal discharge in the initial dewatering stage was determined to be the major cause of cell breakage. The ability of cells to resist breakage was dependant on a combination of factors including host strain, vector, and fermentation strategy. Periplasmic extraction studies were conducted in shake flasks and it was demonstrated that key performance parameters such as Fab titre and nucleic acid concentrations were mimicked. The shake flask system also captured particle aggregation effects seen in a large scale stirred vessel, reproducing the fine particle size distribution that impacts the final centrifugal clarification stage. The use of scale-down Primary Recovery process sequences can be used to screen a larger number of engineered strains. This can lead to closer integration with and better feedback between strain development, fermentation development, and Primary Recovery studies. Biotechnol. Bioeng. 2014;111: 1971–1981. © 2014 Wiley Periodicals, Inc.

  • A mass balance study to assess the extent of contaminant removal achieved in the operations for the Primary Recovery of plasmid DNA from Escherichia coli cells.
    Biotechnology and Bioengineering, 2002
    Co-Authors: Laura A. S. Ciccolini, Parviz Ayazi Shamlou, Nigel J. Titchener-hooker
    Abstract:

    Mass balances were performed on an alkaline lysis operation for the Primary Recovery of supercoiled plasmid DNA as part of a process for plasmid gene preparation. Escherichia coli DH5alpha/pSVbeta was cultured in defined medium by fed-batch fermentation and harvested at the end of the exponential phase. Alkaline lysis of the recombinant cells was performed at fixed shear rates ranging between 46 and 461 s(-1), with neutralization 100 and 300 s after the initiation of the lysis. Mass balance calculations were used to optimize the operating conditions for carrying out the alkaline lysis operation. The results indicated that a plasmid yield of 75% and purity with respect to total DNA of 60% were achievable during the Primary Recovery operation. The influences of key contaminants, including the soluble proteins and the suspended solids, as they bear on the subsequent purification operations, were evaluated and discussed.

Hui Suan Ng - One of the best experts on this subject based on the ideXlab platform.

  • Primary Recovery of recombinant human serum albumin from transgenic Oryza sativa with a single-step aqueous biphasic system
    Journal of The Taiwan Institute of Chemical Engineers, 2018
    Co-Authors: Hui Suan Ng, Chin-chi Wang
    Abstract:

    Abstract Recent advances in genetic engineering technology have provided various methods in recombinant proteins production for pharmaceutical purposes. However, the development of the effective purification strategy remains the major challenge for large-scale production of therapeutic proteins because high purity of the proteins is often demanded. In this study, recombinant Oryza sativa L. cv Tainung 67 human serum albumin (OsrHSA) was expressed and purified with a single-step polyethylene glycol (PEG)/phosphate aqueous biphasic system (ABS). The optimum condition of PEG/phosphate ABS for the purification of OsrHSA was determined by investigating the effects of PEG molecular weight; pH; tie-line length (TLL); and volume ratio (VR) of ABS on the OsrHSA purification efficiency. The purification efficiency of OsrHSA was evaluated based on the partition coefficient (KHSA) and Recovery yield (RB) of the OsrHSA in the PEG/phosphate ABS. An RB of 69.8% of OsrHSA was recovered in the optimum PEG/phosphate ABS consists of TLL of 26.0% (w/w) of pH 7.5. In addition, large-scale ABS were attempted and results showed that the Recovery efficiency of 3000 g ABS was consistent with a 10 g ABS. This novel finding has demonstrated that the feasibility of ABS as a potential tool to purify HSA protein in one-step operation.

  • Primary Recovery of a bacteriocin like inhibitory substance derived from pediococcus acidilactici kp10 by an aqueous two phase system
    Food Chemistry, 2014
    Co-Authors: Sahar Abbasiliasi, Tau Chuan Ling, Shuhaimi Mustafa, Saeid Kadkhodaei, Hui Suan Ng, Tengku Azmi Tengku Ibrahim, Faezeh Vakhshiteh, Zahra Ajdari, Raha Abdul Rahim, Arbakariya B. Ariff
    Abstract:

    A polymer–salt aqueous two-phase system (ATPS) consisting of polyethylene–glycol (PEG) with sodium citrate was developed for direct Recovery of a bacteriocin-like inhibitory substance (BLIS) from a culture of Pediococcus acidilactici Kp10. The influences of phase composition, tie-line length (TLL), volume ratio (VR), crude sample loading, pH and sodium chloride (NaCl) on the partition behaviour of BLIS was investigated. Under optimum conditions of ATPS, the purification of BLIS was achieved at 26.5% PEG (8000)/11% sodium citrate with a TLL of 46.38% (w/w), VR of 1.8, and 1.8% crude load at pH 7 without the presence of NaCl. BLIS from P. acidilactici Kp10 was successfully purified by the ATPS up to 8.43-fold with a yield of 81.18%. Given that the operation of ATPS is simple, environmentally friendly and cost-effective, as it requires only salts and PEG, it may have potential for industrial applications in the Recovery of BLIS from fermentation broth.