Storey B T, Lee C P, Papa S, Rosen S G, Simon G
Biochemistry. 1976 Feb 24;15(4):928-33. doi: 10.1021/bi00649a031.
The intensity of light scattering from suspensions of membrane fragments prepared by sonication of beef heart mitochondria in the presence of EDTA at alkaline pH (ESMP) was determined at 45, 90, and 135 degrees with light of wavelength 546 nm. The dissymmetry ratio Z = I45 degrees c/I135 degrees c, where I45 degrees c and I135 degrees c are the scattering intensities at 45 and 135 degrees extrapolated to zero particle concentration and corrected for reflectance effects, was used to calculate particle size from the Rayleigh-Gans-Debye theory. An average particle diameter D of 184-190 nm was obtained, within the range of particle diameter 50-300 nm determined previously by electron microscopy. This average diameter determined by light scattering is a useful parameter for characterization of ESMP particle size. We propose the term: light scattering average particle diameter, DLS, for this parameter. The refractive index of ESMP was determined to be 1.443 by measurement of scattering intensity in buffer solutions of varying sucrose concentration. The value of Z was independent of sucrose concentration in this determination, showing that the particles are osmotically inactive toward sucrose. The values of average particle diameter DLS and of refractive index fall within the range of validity of the Rayleigh-Gans-Debye theory, for which light scattering changes are attributable solely to dimension change, rather than to change in particle refractive index. Uptake of water accompanying energy-linked salt uptake in ESMP was calculated from light scattering changes to be 0.18 mul of H2O/mg of protein, compared with 0.49 mul of H2O/mg of protein measured by dextran inaccessibility. Measurement of light scattering changes provides a rapid and sensitive method for determining volume changes of ESMP. The magnitude of the volume change observed during energy-linked water and salt uptake and the initial degree of hydration suggests that ESMP are analogous to polyelectrolyte gels with regard to sorption of strong electrolytes and that the Donnan formulation for ion exchange equilibria may be usefully applied to these processes in ESMP.
在碱性pH值下，于存在乙二胺四乙酸（EDTA）的条件下，通过超声处理牛心线粒体所制备的膜片段悬浮液（ESMP）的光散射强度，是在45°、90°和135°下，使用波长为546 nm的光测定的。不对称比率Z = I45°c/I135°c，其中I45°c和I135°c是在45°和135°下外推至零颗粒浓度并校正反射效应后的散射强度，用于根据瑞利 - 甘斯 - 德拜理论计算颗粒大小。获得的平均颗粒直径D为184 - 190 nm，在先前通过电子显微镜确定的50 - 300 nm颗粒直径范围内。通过光散射确定的这个平均直径是表征ESMP颗粒大小的一个有用参数。我们为这个参数提出术语：光散射平均颗粒直径，DLS。通过测量不同蔗糖浓度缓冲溶液中的散射强度，确定ESMP的折射率为1.443。在此测定中，Z值与蔗糖浓度无关，表明颗粒对蔗糖没有渗透活性。平均颗粒直径DLS值和折射率值落在瑞利 - 甘斯 - 德拜理论的有效范围内，对于该理论，光散射变化仅归因于尺寸变化，而非颗粒折射率变化。根据光散射变化计算得出，ESMP中伴随能量相关盐摄取的水摄取量为0.18 μl H₂O/mg蛋白质，相比之下，通过葡聚糖不可及性测量得到的值为0.49 μl H₂O/mg蛋白质。光散射变化的测量为确定ESMP的体积变化提供了一种快速且灵敏的方法。在能量相关的水和盐摄取过程中观察到的体积变化幅度以及初始水合程度表明，就强电解质的吸附而言，ESMP类似于聚电解质凝胶，并且唐南离子交换平衡公式可能可有效地应用于ESMP中的这些过程。
