Protein osmotic pressure determination

Osmotic pressure determinations were made at 27°C on a series of solutions of a globular protein in water. The following results were obtained ... 

A biochemist isolates a new protein and determines its molar mass by osmotic pressure measurements. A 50.0-mL solution is prepared by dissolving 225 mg of the protein in water. The solution has an osmotic pressure of 4.18 mm Hg at 25°C. What is the molar mass of the new protein ... 

Mature human albumin consists of one polypeptide chain of 585 amino acids and contains 17 disulfide bonds. By the use of proteases, albumin can be subdivided into three domains, which have different functions. Albumin has an ellipsoidal shape, which means that it does not increase the viscosity of the plasma as much as an elongated molecule such as fibrinogen does. Because of its relatively low molecular mass (about 69 kDa) and high concentration, albumin is thought to be responsible for 75-80% of the osmotic pressure of human plasma. Electrophoretic smdies have shown that the plasma of certain humans lacks albumin. These subjects are said to exhibit analbuminemia. One cause of this condition is a mutation that affects spUcing. Subjects with analbuminemia show only moderate edema, despite the fact that albumin is the major determinant of plasma osmotic pressure. It is thought that the amounts of the other plasma proteins increase and compensate for the lack of albumin. 

Albumin, which is not glycosylated, is the major protein and is the principal determinant of intravascular osmotic pressure it also binds many Hgands, such as drugs and bilirubin. 

C12-0018. When 7.50 mg of a protein is dissolved in water to give 10.00 mL of solution, the osmotic pressure is found to be 1.66 torr at 21°C. Determine the molar mass of the protein. 

Equation (20-80) requires a mass transfer coefficient k to calculate Cu, and a relation between protein concentration and osmotic pressure. Pure water flux obtained from a plot of flux versus pressure is used to calculate membrane resistance (t ically small). The LMH/psi slope is referred to as the NWP (normal water permeability). The membrane plus fouling resistances are determined after removing the reversible polarization layer through a buffer flush. To illustrate the components of the osmotic flux model. Fig. 20-63 shows flux versus TMP curves corresponding to just the membrane in buffer (Rfouimg = 0, = 0),... 

Figure 15.7 Starling principle a summary of forces determining the bulk flow of fluid across the wall of a capillary. Hydrostatic forces include capillary pressure (Pc) and interstitial fluid pressure (PJ. Capillary pressure pushes fluid out of the capillary. Interstitial fluid pressure is negative and acts as a suction pulling fluid out of the capillary. Osmotic forces include plasma colloid osmotic pressure (np) and interstitial fluid colloid osmotic pressure (n,). These forces are caused by proteins that pull fluid toward them. The sum of these four forces results in net filtration of fluid at the arteriolar end of the capillary (where Pc is high) and net reabsorption of fluid at the venular end of the capillary (where Pc is low).

In order to determine the molecular weight of a particular protein, 0.010 g of the protein was dissolved in water to make 2.93 mL of solution. The osmotic pressure was determined to be 0.821 torr at 20.0°C. What is the molecular weight of the protein ... 

In this equation, u is the osmotic pressure in atmospheres, n is the number of moles of solute, R is the ideal gas constant (0.0821 Latm/K mol), T is the Kelvin temperature, V is the volume of the solution and i is the van t Hoff factor. If one knows the moles of solute and the volume in liters, n/V may be replaced by the molarity, M. It is possible to calculate the molar mass of a solute from osmotic pressure measurements. This is especially useful in the determination of the molar mass of large molecules such as proteins. 

A solution prepared by dissolving 6.95 x 10 1 3 g of protein in 0.0300 L of water has an osmotic pressure of 0.195 torr at 25°C. Assuming the protein is a nonelectrolyte, determine the molar mass of the gene fragment. 

In this form, van t Hoff s law of osmotic pressure is also used to determine the molar masses of biological and synthetic macromolecules. When the osmotic pressure is measured for a solution of macromolecules that contains more than one species of macromolecule (for example, a synthetic pol5mer with a distribution of molar masses or a protein molecule that undergoes association or dissociation), the osmotic pressures of the various solute species II, are additive. That is, in sufficiently dilute solution... 

A student dissolved 25.00 grams of a protein powder that is used as a dietary supplement in water to make exactly 821 mL of solution. The temperature of the solution was determined to be 27°C. He measured the osmotic pressure exerted by the solution and determined it to be 0.0300... 

Alternatively, colloidal plasma expanders (Table 9.3) are used. When administered at appropriate concentrations, they exert an osmotic pressure similar to that of plasma protein, hence vascular volume and blood pressure are maintained. The major disadvantages of colloidal therapy include its relatively high cost, and the risk of prompting a hypersensitivity reaction. Determined elforts to develop blood substitutes were initiated in 1985 by the US military, concerned about the issue of blood supply to future battlefields. 

Because osmotic pressures can be experimentally measured down to rather low values, the Van t Hoff equation proves to be valuable for determining the molecular weights of proteins and other high polymers, as illustrated in Sidebar 7.13. Other practical aspects of osmosis, dialysis, and reverse osmosis phenomena in the physiological and industrial domain are described briefly in Sidebar 7.14. 

Tire evaluation of Mr is often of critical importance. Minimum values of Mr can often be computed from the content of a minor constituent, e.g., the tryptophan of a protein or the iron of hemoglobin. However, physicochemical techniques provide the basis for most measurements.177 Observations of osmotic pressure or light scattering can also be used and provide determinations of Mr that are simple in principle, but which have pitfalls.178... 

Osmotic pressure measurements are particularly convenient for determination of the molar mass of macromolecules such as proteins. 

A solution of crab hemocyanin, a pigmented protein extracted from crabs, was prepared by dissolving 0.750 g in 125 mL of an aqueous medium. An osmotic pressure rise of 2.6 mm of the solution was detected at 4° C. The solution had a density of 1.00 g/mL. Determine the molar mass of the protein. 

Desalting or buffer exchanges are often required between purification steps. At the laboratory scale, the protein solution is placed in a tube of a semipermeable polymer membrane immersed in the desired buffer. The membrane pore size determines the minimum molar mass of the compounds that are retained. Small molecules with a molar mass below the membrane cut-off will flow freely across the membrane until the osmotic pressure equilibrium is reached. Complete buffer exchange requires several changes of the dialysis liquid. The process should be carried out at a temperature around 4°C, to avoid loss of activity. 

To determine the protein concentration Ce in the efferent blood we assume that filtration equilibrium is established before the blood leaves the glomerular capillaries, i.e., that the glomerular hydrostatic pressure minus the efferent colloid osmotic pressure Posm equals the tubular pressure. The experimentally determined relation between the colloid osmotic pressure and the protein concentration C can be described as [16] ... 

The molecular weight of a protein may be determined by the use of thermodynamic methods, such as osmotic pressure measurement, and sedimentation analysis in the ultracentrifuge. Osmotic... 

For a cell membrane in a living animal, a very slight pressure difference will activate transport processes in the membrane, which will effectively eliminate the pressure difference. Introducing this change, there is no longer a state of equilibrium across the membrane, and other transport processes will take place. Such transport often is supported by chemical pumps, which move sodium ions from the protein phase to the aqueous phase. The simple estimation above illustrates that relatively small changes in the concentration are necessary to eliminate the osmotic pressure. In order to force PB — PA = 0, c(Na) in phase B must be reduced by 11 mmol/L, or by less than 10% of the previously determined concentration of 128 mmol/L (Gaiby and Larsen, 1995). 

Figure 9 shows force laws for phosphatidylcholine bilayers (Lis et al, 1982), determined by the osmotic stress method. Similar data were obtained for DNA samples (Rau et al., 1984). The characteristic length governing decay of the force is about 3 A for both systems. Interactions of this kind can also be important for protein aggregates. Prouty et al. (1985) used the osmotic stress method to determine the phase diagram of sickle cell hemoglobin (Fig. 10). At a critical osmotic pressure, which is temperature dependent, a solution of deoxyhemoglobin S collapses to a gel, with a large change in volume. One of the strengths of the osmotic stress method is that it provides additional information that can be used for thermodynamic analysis of the system.

To determine the molar mass of a certain protein, 1.00 X 10 g of the protein was dissolved in enough water to make 1.00 mL of solution. The osmotic pressure of this solution was found to be 1.12 torr at 25.0°C. Calculate the molar mass of the protein. 

Determination of the osmotic pressure of solutions in the presence of other solutes and biologic-binding processes can indicate clearly changes in the amount of water bound or, more exactly, the amount of water that can no longer interact freely with the solutes. Such methods have been useful particularly to determine the DNA-protein interactions (29). 

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