Since the monitoring beam diameter is less than 3 mm, we assume that the I exp value is constant across the whole monitoring beam in the middle of the cuvette. This assumption may not be completely valid for the sample with membranes due to scattering effects. Because of this, scattering effects within membrane bound samples were investigated further. Table 3 Photoexcitation intensities measured at the surface of incidence and estimated at the middle of the sample
cuvette for isolated and membrane-bound RCs 4EGI-1 concentration Parameter I exp at the surface of the cuvette, mW cm−2 Estimated I exp in the middle of the cuvette with isolated RCs, mW cm−2 Estimated I exp in the middle of the cuvette with membrane-bound RCs, mW cm−2 I exp_1 18.07 9.16 0.92 SRT2104 clinical trial I exp_2 9.51 4.82 0.48 I exp_3 7.70 3.91 0.39 I exp_4 5.38 2.76 0.27 I exp_5 3.02 1.52 0.15 I exp_6 AZD8931 ic50 1.59 0.81 0.08 I exp_7 1.29 0.65 0.07 I exp_8 0.69 0.35 0.04 I exp_9 0.39 0.2 0.02 The type and amount of scattering in the membrane samples was estimated by fitting the absorption curve of a membrane sample to the sum of a scaled, previously measured isolated RC absorption spectrum and the scattering formula \( A_\textscatter = C_S \cdot \lambda^K_S \), where C S is a constant and K S characterizes the scattering. For small particles with respect to the wavelength, K S = −4 and is representative of Rayleigh scattering.
Values of K S above −4 and approaching zero are more characteristic of Mie scattering (Cavatorta et al. 1986; Hudson 1969). Figure 5 shows the resulting least squares fit of the membrane absorption spectrum and PI-1840 the corresponding curve for A scatter. From the analysis, the values log[C S ] = 8 ± 0.05 and K S = −2.95 ± 0.02 were obtained. The value of K S indicates that the scattering is more
like that of Mie scattering, or Rayleigh–Debye–Gans scattering, in which case the dimension of the scattering particle was large and could not be treated as a single dipole (Cavatorta et al. 1986; Hudson 1969). The absorption at 802 nm, after subtracting the scatter curve A scatter from the membrane absorption, was used to determine the concentrations to be ~1 µM. This analysis, however, does not address possible multiple scattering effects fully, which were found to play a large role in RC photoexcitation dynamics (Goushcha et al. 2004) and are discussed further below. Fig. 5 Effects of multiple light scattering in membrane-bound RCs. Solid line is membrane absorption curve. Dotted line is the scaled isolated RC spectrum + A scatter. The dashed line below these curves is the contribution due to scattering, A scatter, in the absorption spectrum Figure 6 shows a simplified schematic of the cuvette compartment. The monitoring beam propagates along the x-axis, and CW excitation is applied along the y-axis. Since the scattering is pronounced in membrane samples, the actual CW excitation beam intensity in the middle of the cuvette (the hatched region of the cuvette in Fig.