C. W. Schlapfer and K. Nakamoto vs. time data covering 4 half-lives were in general used and the fit to the equation was in general within the uncertainty of the absorbance measurements.

Incorporation of a biotinylated ruthenium tris(bipyridine) [Ru(bpy) 2 (Biot-bpy)] 2+ (1) in either avidin or streptavidins (strept)avidinscan be conveniently followed by circular dichroism spectroscopy. To determine the stepwise association constants, cooperativity, and chiral discrimination properties, diastereopure (Λ and ∆)-1 species were synthesized and incorporated in tetrameric (strept)avidin to afford (∆-[Ru(bpy) 2 (Biot-bpy)] 2+ ) x ⊂avidin, (Λ-[Ru(bpy) 2 (Biot-bpy)] 2+ ) x ⊂avidin, (∆-[Ru(bpy) 2 (Biot-bpy)] 2+ ) x ⊂streptavidin, and (Λ-[Ru(bpy) 2 (Biotbpy)] 2+ ) x ⊂streptavidin (x ) 1−4) For these four systems, the overall stability constants are log 4 ) 28.6, 30.3, 36.2, and 36.4, respectively. Critical analysis of the CD titrations data suggests a strong cooperativity between the first and the second binding event (x ) 1, 2) and a pronounced difference in affinity between avidin and streptavidin for the dicationic guest 1 as well as modest enantiodiscrimination properties with avidin as host.