Binding properties of 2-acetylnaphthalene with hydroxypropyl cyclodextrins from fluorescence quenching experiments.

The quenching of 2-acetylnaphthalene (2-AN) fluorescence by hydroxypropyl cyclodextrins (HP-CD) has been analyzed using modified Stern-Volmer plots to obtain binding constants as a function of temperature for 2-AN:HP-CD complexes. The HP-CDs were commercially available and contained 4-7 HP groups per CD molecule for alpha-CD, beta-CD, and gamma-CD. HP substitution causes a 12 to over 40% increase in binding constant (K(ave)) for 2-AN compared to that for unsubstituted CDs, although the K(ave) value is not strongly dependent on the extent of HP substitution for beta-CD. No evidence of formation of a 2:2 complex, such as that observed with 2-AN and gamma-CD, is observed with 2-AN and HP-gamma-CD. Thermodynamic parameters (DeltaH degrees and DeltaS degrees ) suggest that the increase in K(ave) with HP substitution is due to an enlarged binding site for the HP-CDs that allows greater motional freedom for 2-AN. Comparison is made to the binding of 2-methylnaphthoate (2-MN) to CDs and HP-CDs, and the larger K(ave) values for 2-MN over 2-AN are attributed to greater dispersion forces for 2-MN complex formation.

The use of neutral cyclodextrins as additives in capillary electrophoresis for the separation and identification of propoxyphene enantiomers.

A simple, efficient, and rapid method is described for separation of the enantiomers of propoxyphene by capillary electrophoresis with neutral cyclodextrins as chiral separators. This method has several advantages over the crystallization method employed by some forensic laboratories, including unambiguous results, ease of use, and smaller sample-size requirement. The method enables baseline separation of the propoxyphene enantiomers in approximately six minutes, which is less than one-third of the time required for a previously published method.

The shape of immunoglobulin G molecules in solution.

We have studied the shape of rabbit Immunoglobulin G molecules in solution by using singlet-singlet energy transfer to determine the minimum distance between the two hapten binding sites. A hybrid antibody was prepared in which one site specifically bound the energy donor, epsilon-dansyl-lysine, and the other site bound the energy acceptor, fluorescein. For this donor-acceptor pair, R(0) was calculated to be 4.8 +/- 0.2 nm (48 +/- 2 A). From a comparison of the lifetime of the donor's excited state in the presence or absence of acceptor, it was found that no energy transfer had occurred in the hybrid. Since the maximum distance over which transfer is measurable was 8.2 nm (82 A; 1.7 R(0)), and since the Fab moieties exhibit segmental flexibility, the average distance between the two hapten-binding sites was estimated to be 9.2-10 nm (92-102 A). If one assumes that the length of the Fab fragment is 7 nm (70 A), the corresponding minimum angle between Fab moieties, alpha(M), would be 80-95 degrees . The molecules in solution, thus, have an open Y- or T-shaped configuration in which the hapten binding sites are not more than 2.5 nm (25 A) from the extreme ends of the Fab fragments. The existence of conformations in which alpha(M) is less than 80 degrees , as has been observed in some antibody-antigen complexes, must therefore be the result of definite conformational changes.