Proteins water adsorption isotherms

A similar roasting effect was shown for the water adsorption isotherms of hull flours except that the most severe roasting did not affect the EMC as dramatically as it did whole flours for the highest aw. The water asorption isotherms of high protein flours indicate that roasting did not affect EMC of these flours... 

Water affects the reaction rate through its effect on reaction kinetics and protein hydration, which is required for optimal enzyme conformation and activity. Enzymes need a small amount of water to maintain their activity however, increasing the water content can decrease the reaction rate as a result of hydrophilic hin-drance/barrier to the hydrophobic substrate, or because of denaturation of the enzyme (189). These opposite effects result in an optimum water content for each enzyme. In SCFs, both the water content of the enzyme support and water solubilized in the supercritical phase determine the enzyme activity. Water content of the enzyme support is, in turn, determined by the distribution/partition of water between the enzyme and solvent, which can be estimated from water adsorption isotherms (141, 152). The solubility of water in the supercritical phase, operating conditions, and composition of the system (i.e., ethanol content) can affect the water distribution and, hence, determine the total amount of water that needs to be introduced into the system to attain the optimum water content of the support. The optimum water content of the enzyme is not affected by the reaction media, as demonstrated by Marty et al. (152), for esterification reaction using immobilized lipase in n-hexane and SCC02- Enzyme activity in different solvents should, thus, be compared at similar water content of the enzyme support. 

For instance, the time course of SPE demonstrates that the solvent phase surfactant concentration steadily decreases (Fig. 3) [58]. The w/o-ME solution s water content decreases at the same rate as the surfactant [58]. The protein concentration at first increases, presumably due to the occurrence of Steps 2 and 3 above, but then decreases due to the adsorption of filled w/o-MEs by the solid phase (Fig. 3) [58]. Additional evidence supporting the mechanism given above is the occurrence of a single Langmuir-type isotherm describing surfactant adsorption in the solid phase for several SPE experiments employing a given protein type (Fig. 4) [58]. Here, solid-phase protein molecules can be considered as surfactant adsorption sites. Similar adsorption isotherms occurred also for water adsorption [58]. 

Hageman et al. [3.13] calculated the absorption isotherms for recombinant bovine somatotropin (rbSt) and found 5-8 g of water in 100 g of protein, which was not only on the surface but also inside the protein molecule. Costantino et al. [3.72] estimated the water monolayer M0 (g/100 g dry protein) for various pharmaceutical proteins and for their combination with 50 wt% trehalose or mannitol as excipient. They compared three methods of calculating MQ (1) theoretical (th) from the strongly water binding residues, (2) from conventional adsorption isotherm measurements (ai) and (3) from gravimetric sorption analysis (gsa) performed with a microbalance in a humidity-controlled atmosphere. Table 3.5 summarizes the results for three proteins. The methods described can be helpful for evaluating RM data in protein formulations. 

Initial high slopes of adsorption isotherms indicate, usually, a high affinity of proteins for the solid/water interfaces (Fig. 6). The AG°ads values calculated from the Langmuir isotherms are usually in the range between —6 and —12 kcal/mol for various protein-adsorbent... 

The protein concentration in the sublayer c(Fi) can be determined via the adsorption isotherm Eqs. (2.117) to (2.119). The Eq. (4.38) is quite complicated for a further analysis and simplifications are necessary. From experimental data, it is known that the adsorption of proteins at the air/water interfaces follows a diffusion-controlled mechanism, at least for small surface pressures n < 2 mN/m [71, 72, 73, 74, 75]. Moreover, the so-called induction time t, the time at which the surface pressure FI starts to increase, can be used for an estimation of the adsorption mechanism. For this time interval the relation cH s const should hold [71, 74, 76]. A diffusion model for the range of small F as approximation was given in [77]... 

A more advanced model was suggested very recently by [78] based on the adsorption isotherm for proteins given by Eq. (2.124). In addition to diffusion of the molecules in the bulk, a kinetic process was assumed equivalent to the mechanism used in the mixed kinetic model. The configuration changes, i.e. orientation of a globular protein molecule to the surface, were characterised by one rate constant k. The following Fig. 4.7 shows model calculations where the following parameters were used coi = 2.5-10 m /mol, W2 = 5.010 m /mol (i.e. coj/ ] = 2), a i = 200. These parameters correspond to those for HSA adsorbed at the water/air interface [79]. The diffusion coefficient was taken to be D = lO cmVs and the protein concentration as 10 mol/I. The equilibrium surface pressure of the protein solutions was taken to be 20 mN/m, typical for HSA at this concentration. It should be noted first that the time required for an experimentally observable decrease of the surface tension, say by 0.5 mN/m, is about 3100 s... 

On the other hand, well-defined adsorption isotherms of proteins have been reported. Figure 1 shows one example, that for chymotrypsin in pure water at 20 C. The attainment of steady surface pressure values, which Increase with increasing protein concentration in solution indicating true equalization of bulk and surface chemical potentials, argues in favor of a reversible adsorption process. In addition, desorption from protein monolayers has been measured. How to rationalize these apparently conflicting results therefore presents an intriguing challenge. 

To illustrate the dependence of the mobility function d>y on the concentration of surfactant in the continuous phase, in Fig. 12 we present theoretical curves, calculated in Ref 138 for the nonionic surfactant Triton X-100, for the ionic surfactant SDS ( + 0.1 M NaCl) and for the protein bovine serum albumin (BSA). The parameter values, used to calculated the curves in Fig. 12, are listed in Table 4 and K are parameters of the Langmuir adsorption isotherm used to describe the dependence of surfactant adsorption, surface tension, and Gibbs elasticity on the surfactant concentration (see Tables 1 and 2). As before, we have used the approximation Dj Dj (surface diffusivity equal to the bulk dif-fusivity). The surfactant concentration in Fig. 12 is scaled with the reference concentration cq, which is also given in Table 4 for Triton X-100 and SDS + 0.1 M NaCl, cq is chosen to coincide with the cmc. The driving force, F, was taken to be the buoyancy force for dodecane drops in water. The surface force is identified with the van der Waals attraction the Hamaker function Ajj(A) was calculated by means of Eq. (86) (see below). The mean drop radius in Fig. 12 is a = 20 /pm. As seen in the figure, for such small drops 4>y = 1 for Triton X-100 and BSA, i.e., the drop sur-... 

The interfacial tension response to transient and harmonic area perturbations yields the dilational rheological parameters of the interfacial layer dilational elasticity and exchange of matter function. The data interpretation with the diffusion-controlled adsorption mechanism based on various adsorption isotherms is demonstrated by a number of experiments, obtained for model surfactants and proteins and also technical surfactants. The application of the Fourier transformation is demonstrated for the analysis of harmonic area changes. The experiments shown are performed at the water/air and water/oil interface and underline the large capacity of the tensiometer. 

Teng, C.D., Zarrintan, M.H., and Groves, M.J. (1991). Water vapor adsorption and desorption isotherms of biologically active proteins. Pharm. Res., 8(2), 191-195. 

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