![Property such as absorbance is proportional to the concentration of complex M An, hence molar ratio of
ligand A and metal M and stability constant can be determined
The equation for complexation can be written as
And the stability constant can be written as
Or in logarithmic form,
Where, k = equilibrium stability constant
[M] = concentration of metal ion uncomplexed
[A] = concentration of ligand (A) uncomplexed
n = Number of moles of ligand combined with one mole of metal ion
[MAn] = concentration of complex](https://image.slidesharecdn.com/complexation3-210713095226/85/Complexation-3-5-320.jpg)
Complexation 3
- 1. UNIT IV COMPLEXATION AND PROTEIN BINDING By: Ms. Swati Gaikwad Assistant Professor Nagpur Pharmacy College ,wanadongri.Nagpur
- 2. METHODS OF ANALYSIS In the study and application of complex compounds, determination of stoichiometric ratio and stability constant is very important. 1. The stoichiometric ratio of ligand-to-metal or donor-to-acceptor. 2. Stability constant of the complex. The general methods and procedure for obtaining these quantity is as follows. 1. Method of continuous variation 2. pH titration method 3. Distribution method 4. Solubility method
- 3. 1. Method of continuous variation • For the measurement of complexation, additive properties such as spectrophotometric extinction coefficient (absorbance), dielectric constant, refractive index etc are used. • If solutions of two species A and B are are mixed & they have no interaction or complexation, then value of property is additive and linear relationship is observed. • If Complexation occurs, the value of additive property will pass through maximum or minimum. • As shown in the figure, dielectric constant is plotted against mole fraction, and a dashed line shows that no complexation occurs and there is a linear relationship, • whereas the break or change in slope in the upper curve indicates formation of complex. • Mole fraction corresponding to the change in slope i.e. 0.5 indicates stoichiometric ratio or 1:1 complex type.
- 4. • When spectrophotometric absorbance is used, observed values at various mole fractions is subtracted from the corresponding values that would have been expected from results of no complex. • The difference D is then plotted against the mole fraction, and from the curve fig ,molar ratio of complex is obtained. • And stability constant can be determined by means of a calculation involving concentration and absorbance difference.
- 5. Property such as absorbance is proportional to the concentration of complex M An, hence molar ratio of ligand A and metal M and stability constant can be determined The equation for complexation can be written as And the stability constant can be written as Or in logarithmic form, Where, k = equilibrium stability constant [M] = concentration of metal ion uncomplexed [A] = concentration of ligand (A) uncomplexed n = Number of moles of ligand combined with one mole of metal ion [MAn] = concentration of complex
- 6. 2. pH titration method • Whenever complexation is accompanied by the change in pH, pH titration method can be used. • It is the most reliable method to be used. • For example, complexation between the cupric ion and Glycine, when Glycine is added to the solution containing the cupric ions, it reacts with cupric ions and release two proton, which decreases the pH of the solution. • The potentiometric titrations are carried out and titration curves are plotted between the pH and equivalents of base added to the solution. • The figure shows that curve obtained with Glycine alone is well above and there is not much change in pH, while the curve for metal Glycine mixture indicates pH is decreased means that the complexation is occurring. • The horizontal distance between the two curves gives the amount of alkali consumed in the reaction, and this quantity is exactly equal to the concentration of ligand bound at that pH.
- 7. Titration curves of cupric-glycine complex, and glycine. the difference in pH for a given quantity of NaoH indicates formation of complex
- 10. Solubility method This method exploits the change in solubility profile as a criterion to analyze the complexation. Whenever the complexation is produced between the two components, there is inhibition or enhancement in the solubility of one of the component. According to the solubility method, in different well stoppered containers solution of complexing agent is placed with excess quantities of drug. These containers agitated in constant temperature bath until equilibrium is attained and supernatant liquid are removed and analyzed. For example, complexation between p-amino benzoic acid and caffeine.
- 11. It is shown in the figure 5.16; the point A crossing the vertical axis indicates the solubility of drug in water. As there is addition of caffeine, there is increase in the solubility of PABA linearly due to complexation. As it reaches point B, there is saturation in the solution and it cannot solubilize drug anymore. Further, increase in caffeine concentration, there is formation of complex and precipitate. At point C all excess PABA passed into the solution and has been converted to complex. But some uncomplexed molecules of PABA combine further with caffeine and form higher complex such as PABA 2- caffeine.
- 12. PROTEIN BINDING Proteins in the body can bind to the drug and can influence drug action in number of ways. Proteins may: • Facilitate the distribution of drugs. • Inactivate the drug by not allowing sufficient concentration of free drug to develop at the receptor site. • Retard the excretion of a drug. • Also cause displacement of body hormones or a co-administered agent. • The structurally altered form of protein is capable of binding with a co- administered drug. • Drug protein complex is biologically active.
- 13. Kinetics of protein binding The interaction between a group or free protein (P) and Drug (D) is written as, The equilibrium constant is Where, (PD) = Concentration of drug protein complex (P) = concentration of unbounded protein (D)= Total concentration of free acid K = Association constant ........................ (2)
- 14. Substituting equation (2) in (1) ……………..…. (3) Let r be the number of moles of drug bound, (PD), per mole of total protein, (Pt); Then Therefore, The ratio r can also be expressed by converting to a linear form, convenient for plotting, by inverting it.
- 15. If ν independent binding sites are available, the expression for r, is simply ν times that for a site, Alternative way to write this equation is to rearrange it first, This is known as scatchard plot.