Separation of albumin

In one of the earlier papers, Sun et al. (48) estimated the binding constants of ibuprofen, flurbiprofen, and ketoprofen to HSA and BSA using the mobility-shift mode of ACE. In this case the drugs were actually used as additives to the background buffer to improve the separation of albumin proteins. This is the opposite approach to what is presented in the next... 

S. Saksena and A.L. Zydney, Effect of solution pH and ionic strength on the separation of albumin from immunoglobulins (IgG) by selective filtration. Biotech. Bioeng. 43 (1994) 960-968. 

El-FFF is a technique devoted to the fractionation of proteins which is reflected in the number of papers applying this technique to protein separations. The possibilities of El-FFF were first demonstrated by Caldwell for the separation of albumin, lysozyme, hemoglobin, and y-globulin in two different buffer solutions (pH 4.5 and 8.0) [35]. Later, the performance of an El-FFF channel with flexible membranes [36], a channel with rigid membranes [256], or a circular channel [260] for the separation of proteins were described. In these studies, human and bovine serum albumin, y-globulin (bovine), cytochrome C (horse heart), lysozyme (egg white) and soluble ribonucleic acid (t-RNA), as well as denaturated proteins, were successfully separated. 

Separation of albumin from globulin, originally done by filtration, was greatly facilitated by Kingsley s introduction of the ether technique (K20). Ether is added to the salt-protein mixture, which is then repeatedly mixed by inversion for 20 seconds and subsequently centrifuged. Flotation of the globulins ensues, and the albumin fraction separates as a subnatant solution. 

Stereo pair TEM showed that the separation of albumin microadsorbates was preserved in surface holes and cracks, in contrast to the Cohn I fibrinogen results. 

Figure 8. The separation of albumin and Ampholine on a 2.5 X 28 cm bed of Sephadex G-50 (fine). Sample volume 6 ml. Eluant 0.1 M phosphate buffer pH 7.

Serum Copper Prior separation of albumin Automatic analyzer (20 per hour) Ashing-1-HNO3 Stopped-flow method CAR ... 

Dmg/plasma protein interactions Asymmetrical Separation of albumin, HDL, a-macroglobulin, and LDL. Determination of drug distribution in FFF fractions using a fluorimetric detector. ... 

Fig. 7. Chromatograms of size-exclusion separation of IgM (mol wt = 800,000) from albumin (69,000) where A—D correspond to IgM aggregates, IgM, monomer units, and albumin, respectively, using (a) FPLC Superose 6 in a 1 x 30 — cm long column, and (b) Sepharose CL-6B in a 37-cm column.

The first observation of the enantioselective properties of an albumin was made in 1958 (28) when it was discovered that the affinity for L-tryptophan exceeded that of the D-enantiomer by a factor of approximately 100. This led to more studies in 1973 of the separation of DL-tryptophan [54-12-6] C22H22N2O2, on BSA immobilized to Sepharose (29). After extensive investigation of the chromatographic behavior of numerous racemic compounds under different mobile-phase conditions, a BSA-SILICA hplc column (Resolvosil-R-BSA, Macherey-Nagel GmvH, Duren, Germany) was... 

The separation of proteins and peptides mixtures is the objective of protein biochemisdy. Albumin (Mr 66 000) concentration in a biological fluid (seaim, urine or cerebrbrospinal fluid) is assayed as markers for a series disease, such as nephritic syndrome or chronic glomuleronephritis. In diabetic patients the progression of microalbuminuria is accompanied by an increase in urinary concentrations of human semm albumen. In normal the excretion of albumin is 20 (tg/ml, in pathology - 20-200 p.g/ml. 

The chromatogram of the protein mixture should show the partial separation of serum albumin and ovalbumin with a trough of at least 30% of height between their peak signals and baseline separation between ovalbumin and cytochrome c. If present in the sample, the dimeric form of serum albumin should also appear as an individual peak signal before elution of the monomeric form. 

Recently, two examples of the separation of enantiomers using CCC have been published (Fig. 1-2). The complete enantiomeric separation of commercial d,l-kynurenine (2) with bovine serum albumin (BSA) as a chiral selector in an aqueous-aqueous polymer phase system was achieved within 3.5 h [128]. Moreover, the chiral resolution of 100 mg of an estrogen receptor partial agonist (7-DMO, 3) was performed using a sulfated (3-cyclodextrin [129, 130], while previous attempts with unsubstituted cyclodextrin were not successful [124]. The same authors described the partial resolution of a glucose-6-phosphatase inhibitor (4) with a Whelk-0 derivative as chiral selector (5) [129]. 

Membranes offer a format for interaction of an analyte with a stationary phase alternative to the familiar column. For certain kinds of separations, particularly preparative separations involving strong adsorption, the membrane format is extremely useful. A 5 x 4 mm hollow-fiber membrane layered with the protein bovine serum albumin was used for the chiral separation of the amino acid tryptophan, with a separation factor of up to 6.6.62 Diethey-laminoethyl-derivatized membrane disks were used for high-speed ion exchange separations of oligonucleotides.63 Sulfonated membranes were used for peptide separations, and reversed-phase separations of peptides, steroids, and aromatic hydrocarbons were accomplished on C18-derivatized membranes. 

Ferric ion was immobilized on a Chelating Sepharose Fast Flow column preparatory to the separation of seven enkephalin-related phosphopep-tides.17 Non-phosphorylated peptides flowed through the column, and the bound fraction contained the product. The capacity of the column was found to be 23 pmol/mL by frontal elution analysis. Cupric ion was immobilized on Chelating Superose for the isolation of bovine serum albumin.18 Cupric ion was immobilized on a Pharmacia HiTrap column for the separation of Protein C from prothrombin, a separation that was used to model the subsequent apparently successful separation of Factor IX from prothrombin Factor IX activity of the eluate was, however, not checked.19 Imidazole was used as the displacement agent to recover p-galactosidase from unclarified homogenates injected onto a nickel-loaded IMAC column.20 Pretreatment with nucleases and cleaning in place between injections were required procedures. A sixfold purification factor was observed. 

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