Spectrophotometric titrations complexes

Spectrophotometric titrations are particularly useful for the analysis of mixtures if a suitable difference in absorbance exists between the analytes and products, or titrant. Eor example, the analysis of a two-component mixture can be accomplished if there is a difference between the absorbance of the two metal-ligand complexes (Eigure 9.33). 

Spectrophotometric titration curve for the complexation titration of a mixture. 

Sketch the spectrophotometric titration curve for the titration of a mixture of 5.00 X 10 M Bi + and 5.00 X 10 M Cu + with 0.0100 M EDTA. Assume that only the Cu +-EDTA complex absorbs at the selected wavelength. 

Data from the spectrophotometric titrations of Fe + with SCN , and of Cu + with EDTA are used to determine the stoichiometry of the resulting complexes using the method of continuous variations. 

Catalytically active species derived from 4. Spectrophotometric titration of the backbone ligand of the sngar discriminating dinuclear copper(ll) complex N, N-bis[(2-pyridylmethyl)-l,3-diaminopropan-2-olato] (//-acetato) dicoppeftll) perchlorate (Cu2(bpdpo), 4) in the presence of two equivalents of copper(ll) ions with sodium hydroxide indicates successive replacement of the bridging acetate anion bound in the sohd state with two hydroxyl ions and two water molecnles in alkaline aqneons solntion (eqs. 2 and 3) (20-22). Two species, [Cu2(L h)(OH)] (4a) and [Cn2(L h)(OH)2] (4b), are thus observed in a pH-dependent equihbrium (20). 

Spontaneous self assembly of a dinuclear triple helical complex is observed with linked bis-[4,5]-pineno-2,2 -bipyridines. Studies by electrospray mass spectrometry, CD and NMR determined that the major species in solution was a complex of Zn L = 2 3 stoichiometry with a triple helical structure and an enantiomerically pure homochiral configuration at the metal centers. The preference for the formation of one of the possible stereoisomers over the other is of interest.265 Another binuclear triple helical complex is formed from zinc addition to bis[5-(l-methyl-2-(6-methyl-2 -pyridyl)benzimidazolyl)]methane. Spectrophotometric titrations with a zinc solution... 

By correlating the observed spectral changes with the concentrations of added cycloamylose, dissociation constants of the cycloamylose-substrate adducts may be calculated (Rossotti and Rossotti, 1961). Values of the dissociation constants determined in this manner for a variety of complexes are presented in Table II. In most cases, stoichiometries of the complexes have been shown to be 1 1 from the presence of distinct isosbestic points in the spectrophotometric titrations. In a few cases, additional spectral perturbations are observed as the cycloamylose concentration is increased, indicating more complex modes of association. Methyl orange, for example,... 

Potentiometric titrations can also be used to determine stability constants for complexes in some cases, which can be useful as a substitute or complementary approach to spectrophotometric titrations (56). By starting at a low pH, where the relative amount of complex formation is low, and then measuring the pH upon addition of standardized base, one can determine the relative amounts of the species in solution or how many equivalents of H+... 

The following example demonstrates the implementation of known spectra and non-absorbing species into the algorithms. It is an aqueous spectrophotometric titration, investigating the complexation of a metal M by a ligand L to form the complex ML. The ligand also acts as a diprotic base and, additionally, the autoprotolysis of the solvent water needs to be taken into account. The complete model is ... 

The product was identihed by a number of spectroscopic methods. Dioxygen uptake was measured by spectrophotometric titration. MALDI-TOF-MS (matrix-assisted laser desorption/ionization-time of flight-mass spectrometry), an MS method particularly suited to determining molecular masses of biopolymers and synthetic materials with relative masses up to several hundred kilodaltons, determined that the product contained stoichiometric amounts of the heme starting material, the copper complex, and dioxygen in a 1 1 1 ratio. 

Although Freudenberg s hypothesis that complex-formation occurred by inclusion within the cavity was generally accepted, there was no direct evidence for this, either in solution or in the solid state. Broser and Lautsch had found by spectrophotometric titration that the complexes of a series of dyes with the cyclodextrins in solution obeyed the mass action law with a stoichiometry of 1 1. They suggested that association on the outside of the ring might not have a defined stoichiometric composition, and they thus interpreted their results as being consistent with inclusion by cyclodextrin. Their results were not conclusive, however. 

The two variables change their role with respect to their dependent versus independent, intensive versus extensive nature. This is also true of e.g. calorimetric, conductometric and spectrophotometric titrations using UV-, IR- or NMR-spectrosco-py We additionally have to consider that in the titration of the catalytic process only the external dynamics are measured a direct comparison with the actual metal fraction of the related intermediate complexes is generally not possible We call this analysis of homogeneous catalytic systems by a metal-ligand titration the method of inverse titration and for the resulting diagrams we use the term li nd-concentration control maps ([L]-control maps) . 

Figure 6.6 Spectrophotometric titration of the binding of inositol hexaphosphate (IHP) to methemoglobin (Methb). The complex has an increased absorbance at 512 and 649 nm, and no increases at 640,618,588, and 599 ran. The concentration of methemoglobin (20 fjM) is about 14 times higher than the dissociation constant of 1.4 f.iM for the complex. The intersection of the slope of the increase in absorbance with the maximum value gives the stoichiometry (1, in this case). Note that this simple procedure cannot be used if the protein is not initially present at such a high concentration relative to the dissociation constant, since the assumption is that all the added ligand is bound to the protein for the early additions.

Fig. 6. Spectrophotometric titration of Co(II) carbonic anhydrase with KCNO. Imidazole-sulfate buffer, pH 6.1, ionic strength 0.07. Enzyme concentration 0,375 mM. The broken curve represents the 1 1 Co(II) enzyme-NCO- complex. Solid curves The visible spectra at 0.84, 0.42 and 0 equivalents of NCO-, respectively...

The interaction of the linearly linked tris-cyclam derivative 13 with Ni(II), Cu(II), Zn(II), Cd(II), and Pd(II) has been investigated [32], As for the above tri-branched systems, all five metals yield solid complexes in which the metal ligand stoichiometry is 3 1 with, for Cu(II), a spectrophotometric titration also confirming the formation of a complex of this stoichiometry in acetonitrile. Cyclic voltammograms of both the Ni(II) (low-spin) and Cu(II) complexes both yield evidence for the presence of M(II)/M(III) as well as M(I)/M(II) couples in acetonitrile. 

The advantage of the kinetic approach to determining acidity and other equilibrium constants lies in the fact that species that do not react with the added substrate do not interfere with the determination. This is in contrast to spectrophotometric titrations, where the change in absorbance with changing pH may be caused by species unrelated to the equilibrium of interest. Such situations are quite common in studies of unstable species encountered in activation of small molecules by transition metal complexes where several species with ionizable hydrogen atoms may coexist in solution. 

EDTA was determined in the urine of patients treated with Na2CaEDTA upon formation of its Fe(II) complex with 2,4,6-tripyridyl-s-triazine at pH 4.5 [30], Concentrations ranging from 0.1-15.7 pM of EDTA in urine could be accurately determined. EDTA can be standardized by spectrophotometric titration with electrolytically pure Cu(II) at pH 5 without an indicator [31]. The break of the titration curve obtained at 700 nm is used as the end-point of the titration. 

A colorimetric method based on the inhibitory effect of EDTA on the Mn(II) catalyzed oxidation of malachite green by periodate was reported [32]. An alternative method based on using Fe(III) instead of Mn(II) was proposed [33]. The reduction of the absorbance of Bi(III) bromo-pyrogallol red tenside ternary complex upon the addition of EDTA has been exploited for its determination. Calibration curves obtained at 650 nm were linear over the range of 0.2-6 pg/mL of EDTA [34]. EDTA in ophthalmic solutions could be assayed by spectrophotometric titration using Mg(II) as the titrant and Arsenazo I as the indicator. The working range was 0.05-2 pg/mL [35]. 

Mixed N- and O-Donor Ligands. Spectrophotometric titration methods have been used to establish the existence and stoicheiometry of V111 complexes with alanine, a-aminobutyric acid, lysine, and glycylglycine pH measurements showed that the amino-acid ligand is co-ordinated in the anion form.385... 

If a molecule has emission properties such as fluorescence or phosphorescence, addition of DNA may, as in the spectrophotometric titrations, result in a batho-chromic shift of the emission maximum. More significantly, the emission intensity may change on complex formation. In the latter circumstances either an increase or a decrease of the emission intensity might be observed [28]. 

Stability constants were determined from spectrophotometric titration data treated by molar-ratio method procedures. In all cases only 1 1 complexes were assumed to form. Thus, the molar absorbtivities and the conditional stability constants in Table IV reflect this assumption. 

The thermodynamics and kinetics of NCS-, CN-,871 Cl-, Br-, NCS- and I- 872 substitutions at [Rh(TPPS)(H2 O)] have been reported the reactions involved are shown in equation (169), and the parameters determined are summarized in Table 60. Spectrophotometric titrations showed two inflection points as OH- is added to [Rh(TPPS)(H20)2]3-, and the consecutive pKx values (7.01 and 9.80 at 20 °C) correspond to the pKk values for fac- and mer-[RhCl3(H20)3], suggesting that the TPPS6- anion and 3 Cl- ligands are comparable electron donors toward the Rh center.872 The trends in the equilibrium constants (Table 60) imply that Rhni is a soft (class B) add in these complexes the NCS- ion is presumed to be sulfur bonded, although no direct evidence is presented to support this assumption. 

An interesting example of difference counting is provided in the conal-bumin study. Conalbumin can bind two iron atoms very tightly and it had been concluded earlier (Warner and Weber, 1953) that each iron atom might be bound to three phenolic groups, which would remain ionized at all pH s where the iron complex is stable. This earlier conclusion was firmly established by spectrophotometric titration of the iron complex. Only five phenolic groups were titrated between pH 8 and 12, compared to eleven in native iron-free conalbumin. The result shows, incidentally. 

The spontaneous assembly of a capped trimetallic complex of type 59 from 57, 58 and copper(I) in acetonitrile-dichloromethane has also been reported. Once again the product was characterised by electrospray mass spectrometry as well as by a spectrophotometric titration study. 

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