Separation of Peptides and Proteins by Countercurrent Chromatography

Xu, Zhongming; Lin, Xianjiang; Lu, Yanbin

doi:10.2174/1570164610666131212000236

Cited by 2 publications

(2 citation statements)

References 0 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…CCC and CPC have numerous advantages such as a high loading capacity and no loss of solute since it is always possible to recover any material trapped in a liquid phase. These advantages were used over the past years to purify biomolecules, especially proteins, by CCC and CPC modes (4,5). Aqueous Two Phase Systems (ATPS) were found very efficient for protein purification (6,7).…”

Section: Introductionmentioning

confidence: 99%

In-situ protein determination to monitor contamination in a centrifugal partition chromatograph

Bouiche¹,

Faure²

2017

Analytical Biochemistry

View full text Add to dashboard Cite

Centrifugal partition chromatography (CPC) works with biphasic liquid systems including aqueous two-phase systems. Metallic rotors are able to retain an aqueous stationary phase able to purify proteins. But the adhesion of proteins to solid surface may pose a cross-contamination risk during downstream processes. So it is of utmost importance to ensure the cleanliness of the equipment and detect possible protein contamination in a timely manner. Thereby, a direct method that allows the determination of the effective presence of proteins and the extent of contamination in the metallic CPC rotors was developed. This in-situ method is derived from the Amino Density Estimation by Colorimetric Assay (ADECA) which is based on the affinity of a dye, Coomassie Brillant Blue (CBB), with protonated N groups of the proteins. In this paper, the ADECA method was developed dynamically, on a 25 mL stainless-steel rotor with various extents of protein contaminations using bovine serum albumin (BSA) as a fouling model. The eluted CBB dye was quantified and found to respond linearly to BSA contamination up to 70 mg injected. Limits of detection and quantification were recorded as 0.9 mg and 3.1 mg, respectively. While the non-specific interactions between the dye and the rotor cannot currently be neglected, this method allows for in situ determination of proteins contamination and should contribute to the development of CPC as a separation tool in protein purification processes.

show abstract

Section: Introductionmentioning

confidence: 99%

In-situ protein determination to monitor contamination in a centrifugal partition chromatograph

Bouiche¹,

Faure²

2017

Analytical Biochemistry

View full text Add to dashboard Cite

show abstract

“…When the method is applied to analytical separation using a narrow bore coiled column, however, the strong cohesive force between the liquid and the tube wall causes the plug flow to lose the stationary phase resulting in the loss of the peak. Consequently, in the past most of protein separations have been performed by semi-preparative to large-scale separations [7–9]. In order to improve the retention of the stationary phase in the analytical column, a spiral tube assembly has been developed [10–13].…”

Section: Introductionmentioning

confidence: 99%

New analytical spiral tube assembly for separation of proteins by counter-current chromatography

Ito

2015

Journal of Chromatography A

View full text Add to dashboard Cite

A new spiral column assembly for analytical separation by counter-current chromatography is described. The column is made from a plastic spiral tube support which has 12 interwoven spiral grooves. The PTFE tubing of 1.6 mm ID was first flattened by extruding through a narrow slit and inserted into the grooves to make 5 spiral layers with about 60 ml capacity. The performance of the spiral column assembly was tested with separation of three stable protein samples including cytochrome C, myoglobin and lysozyme in a polymer phase system composed of polyethylene glycol 1000 and dibasic potassium phosphate each at 12.5 % (w/w) in water. At 2 ml/min, three protein samples were well resolved in one hour. The separation time may be further shortened by application of higher revolution speed and flow rate by improving the strength of the spiral tube support in the future.

show abstract

Separation of Peptides and Proteins by Countercurrent Chromatography

Cited by 2 publications

References 0 publications

In-situ protein determination to monitor contamination in a centrifugal partition chromatograph

In-situ protein determination to monitor contamination in a centrifugal partition chromatograph

New analytical spiral tube assembly for separation of proteins by counter-current chromatography

Contact Info

Product

Resources

About