Analysis of competition experiments

Competition experiments are widely used in the biosciences, particularly in studies of binding interactions. A simple example is shown in Eqn. 9.44. 

In this example, the equilibrium between A, B and AB is affected by the addition of C. The popularity of the competition technique is due to the fact that it can be used to investigate interactions (in this case the binding of A + C = AC) without having to detect the participating species (free C and the complex AC). The method relies on using one interaction (here A + B = AB) as a reporter to monitor the other. 

This assumes, of course, that a suitable signal is available to follow the formation of AB. The diagrams below illustrate (left) the formation of AB, and (right) the effect of adding C to a system containing A, B and AB on addition of C the species AC is formed at the expense of AB whose concentration falls, with a concomitant decrease in the observed signal. 

The analysis of coupled equilibria is the most complex problem that is considered in this chapter. It may seem surprising that such apparently straightforward systems like those shown in Eqns. 9.26 and 9.44 should present such great difficulties in analysis, the more so because it is a trivial matter to calculate the equilibrium constants, if the concentrations of the various species are known. However, that situation arises very rarely for several reasons  

Proceeding as we have done before with complex systems, we calculate theoretical data to deal with these systems. Since straightforward analytical solutions are not available, even for such simple cases as Eqn. 9.44, numerical methods are used to simulate solutions of the equilibria. 

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These data showed that thiamethoxam (13), like imidadoprid (8) and the other neonicotinoids, binds with high affinity to nicotinic receptors [57]. However, there are clear differences to the other commercial neonicotinoids, as documented by a kinetic analysis of competition experiments [56]. While [ H]thiamethoxam (13) binds to receptors with nanomolar affinity, micromolar concentrations are required to displace [ H]imidadoprid (8). Further, the interaction between the two compounds is non-competitive , meaning that binding of thiamethoxam (13) reduces the binding capacity of the receptor preparation for imidadoprid (8) but not its affinity. Thiamethoxam (13) shares this unusual mode of inhibition with other neonicotinoids (not commercialized) containing a N-methyl group as pharmacophore substituent [56, 58]. 

Competitive binding assays are well established in bio-analysis. Over the past decade, substantial efforts have been made to use competition experiments in other research areas, especially supramolecular chemistry. Instead of covering all types of competition experiments, this chapter focuses on a unique competition assay, the so-called indicator displacement assay (IDA), which has become a standard strategy for molecular recognition and sensing. ... 

The competitive adsorption isotherms were determined experimentally for the separation of chiral epoxide enantiomers at 25 °C by the adsorption-desorption method [37]. A mass balance allows the knowledge of the concentration of each component retained in the particle, q, in equilibrium with the feed concentration, < In fact includes both the adsorbed phase concentration and the concentration in the fluid inside pores. This overall retained concentration is used to be consistent with the models presented for the SMB simulations based on homogeneous particles. The bed porosity was taken as = 0.4 since the total porosity was measured as Ej = 0.67 and the particle porosity of microcrystalline cellulose triacetate is p = 0.45 [38]. This procedure provides one point of the adsorption isotherm for each component (Cp q. The determination of the complete isotherm will require a set of experiments using different feed concentrations. To support the measured isotherms, a dynamic method of frontal chromatography is implemented based on the analysis of the response curves to a step change in feed concentration (adsorption) followed by the desorption of the column with pure eluent. It is well known that often the selectivity factor decreases with the increase of the concentration of chiral species and therefore the linear -i- Langmuir competitive isotherm was used ... 

It may be noted that competitive deprotonation of 29 at C-l gives rise to 2-deoxyribonolactone (27) with the concomitant release of free 5-methylcy-tosine as minor processes. Interestingly, competitive hydration of 5-MedCyd radical cations (29) occurs exclusively at C-6 as inferred from labeling experiments with 1802 (36) [61]. Thus, mass spectrometry analysis of the four cis and trans diastereomers of 5-MedCyd glycols 36 showed that incorporation of 1802 takes place exclusively at C-5 of 6-hydroxy-5,6-dihydro-2 -deoxycy-tyd-5-yl radicals (34). 

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