Polarization of the fluorescence of macromolecules. 2. Fluorescent conjugates of ovalbumin and bovine serum albumin

Abstract

POLARIZED FLUORESCENCE OF MACROMOLECULES 155 expected, the first effect is negligible compared to the second, and it is found experimentally that the polarization of the fluorescence from a turbid solution is always lower than one of the same characteristics from a transparent medium.Fortu- nately, the Tyndall effect shown by dilute protein solutions seems to have no detectable effect on the polarization.It is a simple matter to detect in any given case whether depolarization by forward scattering takes place.With the mirror M2 in posi- tion, and the exciting beam traversing the middle of the cell, the average path of the fluorescent light inside the solution is about twice the path in the absence of the mirror.If the polarization of the fluorescence is the same with and without the mirror any depolarization by turbidity can be excluded.SUMMARY 1.A simple addition law for the polarizations of several fluorescent components in solution has been derived.2. It is shown that a system of components which differ in molecular size or in their lifetime of the excited state, but which follow independently Perrin's law of depolarization give in the plot of I/p against Tf/ a curve concave with respect to the latter axis.(p = polarization; T = absolute tempera- ture; X = viscosity of the solvent.)3. An extension of Perrin's theory of depolarization to the case of ellipsoidal molecules carrying randomly oriented oscillators of absorption and emission is described.In such case the polarization of the fluorescence is an explicit function of the principal relaxation times of the rotation of the ellipsoid, the lifetime of the excited state and the limiting polarization.4. A qualitative treatment of the depolarization by intramolecular rotations is given.5. The experimental determination of the polari- zation is described and some causes oferror discussed in detail.