Photophysical Probes of the Amorphous Solid State of Proteins

Abstract

The properties of amorphous solid proteins influence the texture and stability of low-moisture foods, the shelf-life of pharmaceuticals, and the viability of seeds and spores. We have investigated the relationship between molecular mobility and oxygen permeability in dry food protein films-bovine α-lactalbumin (α-La), bovine β-lactoblobulin (β-Lg), bovine serum albumin (BSA), soy 11S globulin, and porcine gelatin-using phosphorescence from the triplet probe erythrosin B. Measurements of the phosphorescence decay in the absence (nitrogen) and presence (air) of oxygen versus temperature provide estimates of the non-radiative decay rate for matrixinduced quenching (k TS0 ) and oxygen quenching (k Q [O 2 ]) of the triplet state. Since the oxygen quenching constant is the product of the oxygen solubility ([O 2 ]) and a term (k Q ) proportional to the oxygen diffusion coefficient, it is a measure of the oxygen permeability through the films. For all proteins except gelatin, Arrhenius plots of k TS0 reveal a gradual increase of apparent activation energy across a broad temperature range starting at ∼50°C; this suggests that there is a steady increase in the available modes of molecular motion with increasing temperature within the protein matrix. Arrhenius plots for k Q [O 2 ] were linear for all proteins with activation energies ranging from 24 to 29 kJ/mol. The magnitude of the oxygen quenching constants varied in the different proteins; the rates were approximately 10-fold higher in α-La, β-Lg, and BSA than in 11S glycinin and gelatin. Although the rate of oxygen permeability was not directly affected by the increased mobility of the protein matrix, plots of k Q [O 2 ] versus k TS0 were linear over nearly three orders of magnitude in the protein films, suggesting that the matrix mobility plays a specific role in modulating oxygen permeability. This effect may reflect differences in matrix-free volume that directly influence both mobility and oxygen solubility.

Revisiting Time-Resolved Protein Phosphorescence

“…1a) were fit to a stretched exponential function. This model has often been used to describe the heterogeneous distribution of relaxation processes in systems such as glasses 12–14 and proteins. 11,15 The fitting parameters for all three temperatures are given in Table I.…”

“…has often been successfully used to describe relaxation processes in glassy or glass-like systems 12–15 and proteins. 11,17–19 In this simple function, I 0 is the initial intensity, τ KWW is the Kohlrausch–Williams–Watts lifetime, and the variable β describes the degree of heterogeneity.…”

“…In addition to MEM-derived lifetime distributions, the stretched exponential function is shown to provide an adequate fit when the phosphorescence signal is due to a single emitter. While many processes—including luminescence decays—that are dependent on relaxation in glassy matrixes have been well described by this simple model, 11–15 we are unaware of any published instances of tryptophan phosphorescence decays fit to a stretched exponential.…”