Ultrafiltration protein

Ultrafiltration Protein removal for plasma sample analysis Molecular-weight-selective membranes Uses special filtration and low-speed centrifuge... 

Low-molecular components were removed from cell-free filtrates of the culture by rotary dialysis and the filtrate was concentrated by ultrafiltration. Proteins were precipitated with (NHi )2S0i (80% saturation) and the precipitate was dissolved and desalted on a Sephadex G-25 column. The protein fraction was concentrated in Aquacid III to a volume of about 1.5 ml which was used as a sample for molecular sieving on a Sephadex G-75 Superfine column (9 x 600 mm). 

Membranes 0.001-0.1 Ultrafiltration protein separation, coneentration and blood fractionation, blood filtering Mesoporous AI2O3, Zr02, Si02... 

Paredes-Lopez and co-workers (1991) extracted protein from a 10% (w/v) solution of defatted chickpea with sodium chloride (0.5 M, pH 7.0) and obtained a chickpea protein isolate containing 87.8% protein using this method. After concentration of the extract by ultrafiltration, protein was flocculated by the addition of water (4°C, pH 7, 1 4 v/v ratio of protein extract water). Mdrquez and co-workers (1996) reported protein contents ranging from 74.7 to 84.2% for common bean protein extract using similar methods. 

PVDF-based microporous filters are in use at wineries, dairies, and electrocoating plants, as well as in water purification, biochemistry, and medical devices. Recently developed nanoselective filtration using PVDF membranes is 10 times more effective than conventional ultrafiltration (UF) for removing vimses from protein products of human or animal cell fermentations (218). PVDF protein-sequencing membranes are suitable for electroblotting procedures in protein research, or for analyzing the phosphoamino content in proteins under acidic and basic conditions or in solvents (219). 

Pish protein concentrate and soy protein concentrate have been used to prepare a low phenylalanine, high tyrosine peptide for use with phenylketonuria patients (150). The process includes pepsin hydrolysis at pH 1.5 ptonase hydrolysis at pH 6.5 to Hberate aromatic amino acids gel filtration on Sephadex G-15 to remove aromatic amino acids incubation with papain and ethyl esters of L-tyrosine and L-tryptophan, ie, plastein synthesis and ultrafiltration (qv). The plastein has a bland taste and odor and does not contain free amino acids. Yields of 69.3 and 60.9% from PPG and soy protein concentrate, respectively, have been attained. 

Fig. 29. Rejection of test proteins as a function of molecular weight, in a series of ultrafiltration membranes with different weight cut-offs (69).

Membrane Sep r tion. The separation of components ofhquid milk products can be accompHshed with semipermeable membranes by either ultrafiltration (qv) or hyperfiltration, also called reverse osmosis (qv) (30). With ultrafiltration (UF) the membrane selectively prevents the passage of large molecules such as protein. In reverse osmosis (RO) different small, low molecular weight molecules are separated. Both procedures require that pressure be maintained and that the energy needed is a cost item. The materials from which the membranes are made are similar for both processes and include cellulose acetate, poly(vinyl chloride), poly(vinyHdene diduoride), nylon, and polyamide (see AFembrane technology). Membranes are commonly used for the concentration of whey and milk for cheesemaking (31). For example, membranes with 100 and 200 p.m are used to obtain a 4 1 reduction of skimmed milk. 

Ultrafiltration. Membranes are used that are capable of selectively passing large molecules (>500 daltons). Pressures of 0.1—1.4 MPa (<200 psi) are exerted over the solution to overcome the osmotic pressure, while providing an adequate dow through the membrane for use. Ultrafiltration (qv) has been particulady successhil for the separation of whey from cheese. It separates protein from lactose and mineral salts, protein being the concentrate. Ultrafiltration is also used to obtain a protein-rich concentrate of skimmed milk from which cheese is made. The whey protein obtained by ultrafiltration is 50—80% protein which can be spray dried. 

Membrane-retained components are collectively called concentrate or retentate. Materials permeating the membrane are called filtrate, ultrafiltrate, or permeate. It is the objective of ultrafiltration to recover or concentrate particular species in the retentate (eg, latex concentration, pigment recovery, protein recovery from cheese and casein wheys, and concentration of proteins for biopharmaceuticals) or to produce a purified permeate (eg, sewage treatment, production of sterile water or antibiotics, etc). Diafiltration is a specific ultrafiltration process in which the retentate is further purified or the permeable sohds are extracted further by the addition of water or, in the case of proteins, buffer to the retentate. 

Electroultrafiltration (EUF) combines forced-flow electrophoresis (see Electroseparations,electrophoresis) with ultrafiltration to control or eliminate the gel-polarization layer (45—47). Suspended colloidal particles have electrophoretic mobilities measured by a zeta potential (see Colloids Elotation). Most naturally occurring suspensoids (eg, clay, PVC latex, and biological systems), emulsions, and protein solutes are negatively charged. Placing an electric field across an ultrafiltration membrane faciUtates transport of retained species away from the membrane surface. Thus, the retention of partially rejected solutes can be dramatically improved (see Electrodialysis). 

Fermentation Processes. The efficient production of penicillin, yeasts, and single-ceUed protein by fermentation requires defoamers to control gas evolution during the reaction. Animal fats such as lard [61789-99-9] were formerly used as a combined defoamer and nutrient, but now more effective proprietary products are usually employed. Defoamer appHcation technology has also improved. For example, in modem yeast production faciHties, the defoamers are introduced by means of automatic electrode-activated devices. One concern in the use of defoamers in fermentation processes is the potential fouHng of membranes during downstream ultrafiltration (qv). SiHcone antifoams (43,44) seem less troubled by this problem than other materials. 

Ultrafiltration (qv) (uf) is increasingly used to remove water, salts, and other low molecular-weight impurities (21) water may be added to wash out impurities, ie, diafiltration. Ultrafiltration is rarely used to fractionate the proteins because the capacity and yield are too low when significant protein separation is achieved. Various vacuum evaporators are used to remove water to 20—40% dry matter. Spray drying is used if a powdery intermediate product is desired. Tyophilization (freeze-drying) is only used for heat-sensitive and highly priced enzymes. 

Membrane Processes Membrane processes are also used diafiltration is convenient for the removal of small contaminating species such as salts and smaller proteins, and can be combined with subsequent steps to concentrate the protein. Provided that proper membrane materials have been selected to avoid protein-membrane interactions, diafiltration using ultrafiltration membranes is typically straightforward, high-yielding and capital-sparing. These operations can often tolerate the concentration or the desired protein to its solu-bihty limit, maximizing process efficiency. 

The largest industrial use of ultrafiltration is the recovery of paint from water-soluble coat bases (primers) applied by the wet electrodeposition process (electrocoating) in auto and appliance factories. Many installations of this type are operating around the world. The recovery of proteins in cheese whey (a waste from cheese processing) for dairy applications is the second largest application, where a... 

Van den Berg, G.B. Hanemajer, J.H. and Smolders, C.A., "Ultrafiltration of Protein Solutions the Role of Protein Association in Rejection and Osmotic Pressure," Journal of Membrane Science, 31 (1987) 307-320. 

Ultrafiltration is an improvement on the dialysis principle. Filters having pore sizes over the range of biomolecular dimensions are used to filter solutions to select for molecules in a particular size range. Because the pore sizes in these filters are microscopic, high pressures are often required to force the solution through the filter. This technique is useful for concentrating dilute solutions of macromolecules. The concentrated protein can then be diluted into the solution of choice. 

Ultrafiltration of micellar solutions combines the high permeate flows commonly found in ultrafiltration systems with the possibility of removing molecules independent of their size, since micelles can specifically solubilize or bind low molecular weight components. Characteristics of this separation technique, known as micellar-enhanced ultrafiltration (MEUF), are that micelles bind specific compounds and subsequent ultrafiltration separates the surrounding aqueous phase from the micelles [70]. The pore size of the UF membrane must be chosen such, that the micelles are retained but the unbound components can pass the membrane freely. Alternatively, proteins such as BSA have been used in stead of micelles to obtain similar enan-tioselective aggregates [71]. 

Ultrafiltration Pressure gradient <0.1 p.m-5 nm Emulsions, colloids, macromolecules, proteins... 

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