Protein separation and

Economic Yield Both in a high-value protein separation and in a low-value commodity concentration, economic yield is vital. Economic yield is defined as the fraction of useful product entering the process that leaves it in salable form. The yield equations used in the industry focus on retention, so they deal only with direct losses through the membrane. These losses result both in direct (product not sold) and indirect costs from a waste stream whose disposal or subsequent use may be more expensive when it is contaminated by macrosolute. There are additional indirec t losses, mainly product left in the equipment, particularly that left adhering to the membrane. Costs of cleaning and disposal or this indirect loss, while hard to measure, are usually higher than the cost of product lost through the membrane. 

Extensive research has been carried out into the molecular aspects of foreign protein production in whole plants to enhance the yield, quality and stability of the product and to facilitate protein separation and purification from the biomass [3, 6, 9], In contrast, comparatively little research has been undertaken to investigate the... 

Figure 7.7. Results from model simulations showing the effect of protein separation and the effect of MS detection limit and MS dynamic range on the success rate and the relative dynamic range (RDR) for detection of proteins from H. sapiens tissue samples. (See page 219 for text discussion.)...

One of the major advantages of CE as a separation technique is the wide variety of separation modes available. Analytes can be separated on the basis of charge, molecular size or shape, pi, or hydrophobicity. The same CE instrument can be used for zone electrophoresis, IEF, sieving separations, isotachophoresis, and chromatographic techniques such as MEKC and capillary electrokinetic chromatography. This section provides a brief description of each separation mode. Zone electrophoresis, IEF, and sieving are the primary modes used for protein separations, and these will be discussed in detail in the following sections. 

Proteins can be separated using the 2D electrophoresis method. The first dimension is separation according to the pH of the proteins. The proteins are placed on a gel strip in a buffer solution. An electrical current is applied and the proteins separate and migrate to their isoelectric points (pl)-... 

P Sun, A Hoops, RA Hartwick. Enhanced albumin protein separations and protein-drug binding constant measurements using anti-inflammatory drugs as run buffer additives in affinity capillary elctrophoresis. J Chromatogr B 661 335-340, 1994. 

The nomenclature of the RP is not consequent. The RP most often used contains octyl (RP C8) or octadecyl (RP C18) groups. There is no differentiation even when two methyl groups are introduced additionally with the silane (as with monofunctional silanes) or only one (difunctional) or none (trifunctional silane). Some manufacturer use silanes with bulky side groups (e.g., isopropyl groups) to improve the hydrolytic stability of the bonded phases, but here also, only the longest alkyl group is used in nomenclature. RP C8 and RP C18 are the work horses in HPLC. Shorter chains (RP4) are used in protein separations, and special selectivity can be obtained with bonded phenyl, cyano, amino or fluoro groups. 

Most of the applications of HPLC for protein analysis deal with the storage proteins in cereals (wheat, corn, rice, oat, barley) and beans (pea, soybeans). HPLC has proved useful for cultivar identihcation, protein separation, and characterization to detect adulterations (illegal addition of common wheat flour to durum wheat flour) [107]. Recently Losso et al. [146] have reported a rapid method for rice prolamin separation by perfusion chromatography on a RP POROS RH/2 column (UV detection at 230nm), sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis (PAGE), and molecular size determination by MALDl-MS. DuPont et al. [147] used a combination of RP-HPLC and SDS-PAGE to determine the composition of wheat flour proteins previously fractionated by sequential extraction. 

Proteome analysis in general involves two stages protein separation and subsequent identification and analysis. Multidimensional separations are required in order to result in an adequate resolution of complex protein or peptide... 

At about the same time, the field of protein separation and purification was undergoing rapid development. The introduction of materials for protein chromatography, such as cross-linked dextran, agarose, and polyacrylamide, provided a means to study protein-surface interactions, as well as to dramatically advance knowledge in protein biochemistry. 

It is one of the most effective methods of protein separation and characterization. The chief advantages of this method are that it can be performed under very mild conditions and it has high resolving power, resulting in the clear separation of similarly charged protein molecules. However, in order to use this separation technique, the components of a mixture must have an ionic form, and each component must possess a different net charge. 

Research on microchip protein analysis has been very active for cellular protein functional assay, clinical diagnostics, and proteomics studies. Once again, the microfluidic technology plays an important role in protein assays. Immunoassay, protein separation, and enzymatic assay will be described in detail in subsequent sections. 

Flow Field-Flow Fractionation as a Methodology for Protein Separation and Characterization, J. C. Giddings, F. J. Yang, and M. N. Myers, Anal. Chem., 81, 395... 

With these newer methods of protein separation and amino acid analysis he prepared serum protein fractions by serial salting out with ammonium sulfate and by the Sober and Peterson DEAE cellulose columns (42), using the sera of reptile, fowl, and mammalian blood. Some of the amino acid analyses were carried out by the automatic amino acid methods of Hirs, Moore, and Stein (18). Fortified with this plethora of data, Block now had the opportunity to re-examine not only the ratio of the basic amino acids, but at least 12 amino acids in a variety of protein fractions prepared by at least two different procedures. With the aid of a statistician he determined the significance of the constancy of the molar ratios of pairs of amino acids and found that in spite of the marked variation of the absolute amounts of an amino acid, the molar ratios of certain pairs remain relatively constant among the numerous protein components of animal sera. 

Shortcomings in current understanding of the thermodynamics of protein solutions can be illustrated by considering one of the oldest protein concentration methods, precipitation, and one of the newest and most technologically interesting methods for protein separation and purification, aqueous... 

Chapter 15 focuses mainly on antibody purification by chromatographic means. Numerous sorbents have been developed for protein separation, and they are based on a variety of adsorption-desorption principles. Selection of suitable materials and principles depends on the properties of the particular immunoglobulins to be separated and on the composition of the impurities that constitute the feedstock. 

If the sample is readily soluble in the mobile phase, GPC is unmatched by any other mode of chromatography for simplicity, since the entire analysis is accomplished in a column volume. The time and effort required to develop a separation is less than any other mode of HPLC. It can be of immense value in the purification or organic and inorganic synthesis reaction mixtures, purification of natural products extracts, and for the rapid clean-up of extracts (from plants, insects, soil, etc.) prior to the assay of small molecules. Aqueous size separation is referred to as gel filtration chromatography and is very useful for protein separations and the analysis of water-soluble polymers. 

Badal MY, Wong M, Chiem N, Salimi-Moosavi H, Harrison DJ. Protein separation and surfactant control of electroosmotic flow in poly(dimethylsiloxane)-coated capillaries and microchips. J Chromatogr A 2002 947 277-286. 

The development of novel supports and stationary phases, in particular for protein separation, and the understanding of the separation mechanisms are still in their infancy further improvements must be made. 

Interactions between protein and polysaccharide have attracted increasing interests in the past two decades also because of their implications in many biological processes such as the organization of living cells and the use in many industrial applications such as microencapsulation, protein separation and purification, and in processed foods (Berdick and Morawetz 1954 Burgess and Carless 1984 Dubin et al. 1994 Tolstoguzov 1991). 

In these methods, the separation is carried out by a chromatographic setup. This setup may use various chromatographic principles for protein separation, and this... 

Jun BH, Noh MS, Kim G, Kang H, Kim JH, Chung WJ, Kim MS, Kim YK, Cho MH, Jeong DH, Lee YS (2009) Protein separation and identification using magnetic beads encoded with surface-enhanced Raman spectroscopy. Anal Biochem 391 24—30... 

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