Purification of chymotrypsin from bovine pancreas using precipitation with a strong anionic polyelectrolyte

Boeris, Valeria; Romanini, Diana; Farruggia, Beatriz; Picó, Guillemo

doi:10.1016/j.procbio.2009.02.009

Cited by 31 publications

(34 citation statements)

References 10 publications

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“…In the precipitation process, poly-charged molecules which contain opposite electrical charges to the chymotrypsinogen are added into protein solutions to form a chymotrypsinoge n-polyelectrolyte complex and generate insoluble aggregates. The materials used in the process are salts, non-ionicpolymers, organic solvents and polyelectrolytes which are easy to bioseparate in the downstream process (Boeris et al, 2009;Chatterjeeet al, 2004). Yang (2009) used ammonium sulfate saturation 30-60% in chymotrypsin extraction; Kristjansson and Nielson (1992), Castillo-Yañez et al (2006 and Heu (1995) used ammomium sulfate saturation 30-70% and the recovery yield were in the rang from 55-60%.…”

Section: Ammonium Sulfate Methodmentioning

confidence: 99%

“…Yang (2009) used ammonium sulfate saturation 30-60% in chymotrypsin extraction; Kristjansson and Nielson (1992), Castillo-Yañez et al (2006 and Heu (1995) used ammomium sulfate saturation 30-70% and the recovery yield were in the rang from 55-60%. Boeris (2009) used Polyvinyl Sulfonate (PVS) as polyelectrolytes during the purification process to produce a chymotrypsinogen-PVS precipitate complex in acid pH which decreased the recovery rate of chymotrypsin. Also, increasing the PVS concentration lead to a lower recovery rate but produced a higher purity.…”

Section: Ammonium Sulfate Methodmentioning

confidence: 99%

“…The inhibitors in blood can reduce chymotrypsin activity (Chong et al, 2001;Boeris et al, 2009). The intestine is then chopped into small pieces and mixed with 50 mM Tris-HCl buffer having a pH of 7.5.…”

Section: Extractionmentioning

confidence: 99%

“…Precipitation is the technique most widely used in the chymotrypsin production industry because it can effectively purify and concentrate chymotrypsin at a very low cost (Matsudo et al, 2003;Boeris et al, 2009).Generally, chymotrypsin precipitation is used to obtain crude chymotrypsinogen and can be combined with chymotography techniques for further purification. Purification can be accomplished either by precipitation (using Tris-HCl) and ammonium sulfate fractionation or formation of reverse micelles (Mockel and Barnard, 1969;Simpson, 2000;Wilk, 2001;.…”

Section: Purificationmentioning

confidence: 99%

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Extraction, Purification and Characterization of Fish Chymotrypsin: A Review

K.¹

2011

American Journal of Biochemistry and Biotechnology

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Problem statement: Solid fish waste is generated from the unwanted parts of fish including heads, tails, fins, frames, offal (guts, kidney and liver) and skin. It accounts for up to 80% of material from production of surimi, 66% from production of fillet and 27% from production of headed and gutted fish. Currently, fish wastes are disposed off in land-based waste disposal systems or at sea generating toxic byproducts during the decomposition process. However, fish processing waste can be used to produce commercially valuable by-products, such as chymotrypsin. Approach: A comperehensive review of the literature on the extraction, purification and characeterization of fish chymotrypsin was performed. Results: Chymotrypsin is an endopeptidase secreted by the pancreatic tissues of vertebrates and invertebrates. It has 3 different structures (chymotrypsin A, B and C) varying slightly in solubility, electrophoretic mobility, isoelectric point and cleavage specificity. Only chymotrypsin A and B are found in fish. Compared with mammal chymotrypsin, fish chymotrypsins have similar amino acid composition and molecular weights. Fish chymotrypsins have higher specific activity, especially those from cold-water fish, and low pH and temperature tolerance. The factors affecting the concentration and activity of chymotrypsin in fish are water temperature, fish species, fish age, fish weight and starvation. Chymotrypsin has application in various industries including the food industry, leather production industry chemical industry and medical industry. Conclusion: Extraction techniques for chymotrypsin include: ultra-filtration, ammonium sulphate fractionation precipitation or water-in-oil microemulsions. Purification can be carried out using recrystallization and gel-filtration, ion-exchange and hydrophobic interaction chromatography. Further studies should focus on the optimization of purifiying chymotrypsin from fish processing wastes.

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Section: Ammonium Sulfate Methodmentioning

confidence: 99%

Section: Ammonium Sulfate Methodmentioning

confidence: 99%

Section: Extractionmentioning

confidence: 99%

Section: Purificationmentioning

confidence: 99%

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Extraction, Purification and Characterization of Fish Chymotrypsin: A Review

K.¹

2011

American Journal of Biochemistry and Biotechnology

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“…This technique offers the possibility of concentrating and purifying the target macromolecule usually at a low cost (Boeris et al 2009, Soares et al 2012. Precipitation should result both in concentration and in purification, but actually the former is more effectively achieved.…”

mentioning

confidence: 99%

Purification of inulinases by changing the ionic strength of the medium and precipitation with alcohols

Golunski

Silva

Marques

et al. 2017

An. Acad. Bras. Ciênc.

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The present study evaluated the purification of inulinase by changing the ionic strength of the medium by addition of NaCl and CaCl 2 followed by precipitation with n-propyl alcohol or iso-propyl alcohol. The effects of the concentration of alcohols and the rate of addition of alcohols in the crude extract on the purification yield and purification factor were evaluated. Precipitation caused an activation of enzyme and allowed purification factors up to 2.4-fold for both alcohols. The purification factor was affected positively by the modification of the ionic strength of the medium to 0.5 mol.L -1 NaCl before precipitation with the alcohol (n-propyl or iso-propyl). A purification factor of 4.8-fold and an enzyme yield of 78.1 % could be achieved by the addition of 0.5 mol.L -1 of NaCl to the crude extract, followed by the precipitation with 50 % (v/v) of n-propyl alcohol, added at a flow rate of 19.9 mL/min.

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Customization of copolymers to optimize selectivity and yield in polymer‐driven antibody purification processes

Capito

Skudas

Stanislawski

et al. 2013

Biotechnology Progress

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This manuscript describes customization of copolymers to be used for polymer-driven protein purification in bioprocessing. To understand how copolymer customization can be used for fine-tuning, precipitation behavior was analyzed for five target antibodies (mAbs) and BSA as model impurity protein, at ionic strength similar to undiluted cell culture fluid. In contrast to the use of standardized homopolymers, customized copolymers, composed of 2-acrylamido-2-methylpropane sulfonic acid (AMPS) and 4-(acryloylamino)benzoic acid (ABZ), exhibited antibody precipitation yields exceeding 90%. Additionally, copolymer average molecular weight (Mw ) was varied and its influence on precipitation yield and contaminant coprecipitation was investigated. Results revealed copolymer composition as the major driving force for precipitation selectivity, which was also dependent on protein hydrophobicity. By adjusting ABZ content and Mw of the precipitant for each of the mAbs, conditions were found that allowed for high precipitation yield and selectivity. These findings may open up new avenues for using polymers in antibody purification processes.

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Purification of chymotrypsin from bovine pancreas using precipitation with a strong anionic polyelectrolyte

Cited by 31 publications

References 10 publications

Extraction, Purification and Characterization of Fish Chymotrypsin: A Review

Extraction, Purification and Characterization of Fish Chymotrypsin: A Review

Purification of inulinases by changing the ionic strength of the medium and precipitation with alcohols

Customization of copolymers to optimize selectivity and yield in polymer‐driven antibody purification processes

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