• EDTA Titrations

• Chelation in Biochemistry

• Metal-Chelate Complexes

• Metals are Lewis acids that accept electron pairs from donating ligands that act as Lewis bases

–CN- is a common monodentate ligand, binding to a metal ion through one atom (C)

–Metals can bind to multiple ligands (usually 6)

• A ligand that can attach to a metal by more than one atom is multidentate or a chelating ligand

• Chelating agents can be used for titration of metals to form complex ions (complexometric titration)

• Chelating Agents in Analytical Chemistry

• Ethylenediamenetetraacetic acid (EDTA)

• Acid/Base Properties of EDTA

• EDTA is a hexaprotic system (H6Y2+) with 4 carboxylic acids and 2 ammoniums:

• We usually express the equilibrium for the formation of complex ion in terms of the Y4- form (all six protons dissociated). You should not take this to mean that only the Y4- form reacts

• Fraction of EDTA in Y4- Form

• Similar to acids and bases, we can define fractional compositions, α, defined as the fraction of “free” EDTA in a particular form.

–“Free” means uncomplexed EDTA

–So, for Y4-:

• EDTA Complexes

• The equilibrium constant for a reaction of metal with EDTA is called the formation constant, Kf, or the stability constant:

• Again, Kf could have been defined for any form of EDTA, it should not be understood that only the Y4- reacts to form complex ion.

• pH Dependence of αY4-

• Formation Constants for M-EDTA Complexes

• Some Metals Form 7 or 8 Coordinate Complexes

• Conditional Formation Constant

• We saw from the fraction plot that most of the EDTA is not in the form of Y4- below a pH ~10.

• We can derive a more useful equilibrium equation by rearranging the fraction relationship:

• If we fix the pH of the titration with a buffer, then αY4- is a constant that can be combined with Kf

• Example

• Calculate the concentration of free Ca2+ in a solution of 0.10 M CaY2- at pH 10 and pH 6. Kf for CaY2- is 4.9×1010 (Table 13-2)

• At low pH, the metal-complex is less stable

• Calcium/EDTA Titration Curve

• Generic Titration Curve

• Before the Equivalence Point

• What’s pCa2+ when we have added 5.0 mL of EDTA?

• At the Equivalence Point

• What’s pCa2+ when we have added 25.0 mL of EDTA?

–At the equivalence point almost all the metal is in the form CaY2-

–Free Calcium is small and can be found w/ algebra

• After the Equivalence Point

• What’s pCa2+ when we have added 26.0 mL of EDTA?

–We have 1.0 mL excess EDTA

• Auxiliary Complexing Agents

• In aqueous solution, metal-hydroxide complexes or precipitates can form, especially at alkaline pH

• We often have to use an auxiliary complexing agent

–This is a ligand that binds strongly enough to the metal to prevent hydroxide precipitation, but weak enough to be displaced by EDTA

• Ammonia is a common auxiliary complex for transition metals like zinc

• Metal Ion Indicators

• To detect the end point of EDTA titrations, we usually use a metal ion indicator or an ion-selective electrode (Ch. 15)

• Metal ion indicators change color when the metal ion is bound to EDTA:

–Eriochrome black T is an organic ion

• The indicator must bind less strongly than EDTA

• Metal Ion Indicator Compounds

• EDTA Titration Techniques

• Direct titration: analyte is titrated with standard EDTA with solution buffered at a pH where Kf’ is large

• Back titration: known excess of EDTA is added to analyte. Excess EDTA is titrated with 2nd metal ion.

• EDTA Titration Techniques (2)

• Displacement titration: For metals without a good indicator ion, the analyte can be treated with excess Mg(EDTA)2-. The analyte displaces Mg, and than Mg can be titrated with standard EDTA

• Indirect titration: Anions can be analyzed by precipitation with excess metal ion and then titration of the metal in the dissolved precipitate with EDTA.

• Example Titration

• 25.0 mL of an unknown Ni2+ solution was treated with 25.00 mL of 0.05283 M Na2EDTA. The pH of the solution was buffered to 5.5 and than back-titrated with 17.61 mL of 0.02299 M Zn2+. What was the unknown Ni2+ M?

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EDTA Titrations
Introduction

1.) Metal Chelate Complexes
Any reagent which reacts with an analyte in a known ratio and with a large equilibrium constant can potentially be used in a titration.

Complexation Titrations are based on the reaction of a metal ion with a chemical agent to form a metal-ligand complex.
Metal
Ligand
Metal-Ligand Complex
Metal – Lewis Acid or Electron-pair acceptor
Ligand – Lewis Base or Electron-pair donor
Note: multiple atoms from EDTA are binding Mn2+

EDTA Titrations
Introduction

1.) Metal Chelate Complexes
Complexation Titrations are essentially a Lewis acid-base reaction, in which an electron pair is donated from one chemical to another
The ligands used in complexometric titrations are also known as chelating agents.
Ligand that attaches to a metal ion through more than one ligand atom
Most chelating agents contain N or O
Elements that contain free electron pairs that may be donated to a metal
Fe-DTPA Complex

EDTA Titrations
Metal Chelation in Nature

1.) Potassium Ion Channels in Cell Membranes
Electrical signals are essential for life
Electrical signals are highly controlled by the selective passage of ions across cellular membranes
Ion channels control this function
Potassium ion channels are the largest and most diverse group
Used in brain, heart and nervous system
Current Opinion in Structural Biology 2001, 11:408–414
Opening of potassium channel allows K+ to exit cell and change the electrical potential across membrane
K+ channel spans membrane
channel contains pore that only allows K+ to pass
K+ is chelated by O in channel
http://www.bimcore.emory.edu/home/molmod/Wthiel/Kchannel.html

EDTA Titrations
Metal –Chelate Complexes

1.) Formation Constant (Kf)
The equilibrium constant for the reaction between a metal ion (M+n) and a chelating agent (L-P) is known as a formation constant or stability constant.

Applying different and specific names to the general equilibrium constant is a common occurrence
Solubility (Ksp), acid-base (Ka, Kb), water dissociation (Kw), etc

Chelate effect: ability of multidentate ligands to form stronger metal complexes compared to monodentate ligands.
Kf = 8×109
Kf = 4×109
2 ethylenediamine molecules binds tighter than 4 methylamine molecules

EDTA Titrations
Metal –Chelate Complexes

2.) Chelate Effect
Usually chelating agents with more than one electron pair to donate will form stronger complexes with metal ions than chelating agents with only one electron pair.
Typically more than one O or N
Larger Kf values

Multidentate ligand: a chelating agent with more than one free electron pair
Stoichiometry is 1:1 regardless of the ion charge

Monodentate ligand: a chelating agent with only one pair of free electrons
Multidentate ligand that binds radioactive metal attached to monoclonal antibody (mAb).

mAb is a protein that binds to a specific feature on a tumor cell delivering toxic dose of radiation.

EDTA Titrations
EDTA

1.) EDTA (Ethylenediaminetetraacetic acid)
One of the most common chelating agents used for complexometric titrations in analytical chemistry.

EDTA has 6 nitrogens & oxygens in its structure giving it 6 free electron pairs that it can donate to metal ions.
High Kf values
6 acid-base sites in its structure

EDTA Titrations
EDTA

2.) Acid-Base Forms
EDTA exists in up to 7 different acid-base forms depending on the solution pH.

The most basic form (Y4-) is the one which primarily reacts with metal ions.
EDTA-Mn Complex

EDTA Titrations
EDTA

2.) Acid-Base Forms
Fraction (α) of the most basic form of EDTA (Y4-) is defined by the H+ concentration and acid-base equilibrium constants
Fraction (α) of EDTA in the form Y4-:
where [EDTA] is the total concentration of all free EDTA species in solution
αY4- is depended on the pH of the solution

EDTA Titrations
EDTA

3.) EDTA Complexes
The basic form of EDTA (Y4-) reacts with most metal ions to form a 1:1 complex.
Other forms of EDTA will also chelate metal ions

Recall: the concentration of Y4- and the total concentration of EDTA is solution [EDTA] are related as follows:
Note: This reaction only involves Y4-, but not the other forms of EDTA
where αY4-is dependent on pH

EDTA Titrations
EDTA

3.) EDTA Complexes
The basic form of EDTA (Y4-) reacts with most metal ions to form a 1:1 complex.

EDTA Titrations
EDTA

3.) EDTA Complexes
Substitute [Y4-] into Kf equation

If pH is fixed by a buffer, then αY4- is a constant that can be combined with Kf
where [EDTA] is the total concentration of EDTA added to the solution not bound to metal ions
Conditional or effective formation constant:
(at a given pH)

EDTA Titrations
EDTA

3.) EDTA Complexes
Assumes the uncomplexed EDTA were all in one form
at any pH, we can find αY4- and evaluate Kf’

EDTA Titrations
EDTA

4.) Example:
What is the concentration of free Fe3+ in a solution of 0.10 M Fe(EDTA)- at pH 8.00?

EDTA Titrations
EDTA

5.) pH Limitation
Note that the metal –EDTA complex becomes less stable as pH decreases
Kf decreases
[Fe3+] = 5.4×10-7 at pH 2.0 -> [Fe3+] = 1.4×10-12 at pH 8.0

In order to get a “complete” titration (Kf ≥106), EDTA requires a certain minimum pH for the titration of each metal ion
End Point becomes less distinct as pH is lowered, limiting the utility of EDTA as a titrant

EDTA Titrations
EDTA

5.) pH Limitation
By adjusting the pH of an EDTA titration:
one type of metal ion (e.g. Fe3+) can be titrated without interference from others (e.g. Ca2+)
Minimum pH for Effective Titration of Metal Ions

EDTA Titrations
EDTA Titration Curves

1.) Titration Curve
The titration of a metal ion with EDTA is similar to the titration of a strong acid (M+) with a weak base (EDTA)

The Titration Curve has three distinct regions:
Before the equivalence point (excess Mn+)

At the equivalence point ([EDTA]=[Mn+]

After the equivalence point (excess EDTA)

EDTA Titrations
EDTA Titration Curves

2.) Example
What is the value of [Mn+] and pM for 50.0 ml of a 0.0500 M Mg2+ solution buffered at pH 10.00 and titrated with 0.0500 m EDTA when (a) 5.0 mL, (b) 50.0 mL and (c) 51.0 mL EDTA is added?
Kf = 108.79 = 6.2×108
αY4- at pH 10.0 = 0.30
mL EDTA at equivalence point:
mmol of EDTA
mmol of Mg2+

EDTA Titrations
EDTA Titration Curves

2.) Example
(a) Before Equivalence Point ( 5.0 mL of EDTA)
Before the equivalence point, the [Mn+] is equal to the concentration of excess unreacted Mn+. Dissociation of MYn-4 is negligible.
moles of Mg2+
originally present
moles of EDTA added
Original volume
solution
Volume titrant
added
Dilution effect

EDTA Titrations
EDTA Titration Curves

2.) Example
(b) At Equivalence Point ( 50.0 mL of EDTA)
Virtually all of the metal ion is now in the form MgY2-
Original [Mn+] Original volume of
Mn+ solution
Original volume
solution
Volume titrant
added
Dilution effect
Moles Mg+ ≡ moles MgY2-

EDTA Titrations
EDTA Titration Curves

2.) Example
(b) At Equivalence Point ( 50.0 mL of EDTA)
The concentration of free Mg2+ is then calculated as follows:
Initial Concentration (M)
0
0
0.0250
Final Concentration (M)
x
x
0.0250 – x
Solve for x using the quadratic equation:

EDTA Titrations
EDTA Titration Curves

2.) Example
(c) After the Equivalence Point ( 51.0 mL of EDTA)
Virtually all of the metal ion is now in the form MgY2- and there is excess, unreacted EDTA. A small amount of free Mn+ exists in equilibrium with MgY4- and EDTA.
Original [EDTA] Volume excess
titrant
Original volume
solution
Volume titrant
added
Dilution effect
Excess moles EDTA
Calculate excess [EDTA]:

EDTA Titrations
EDTA Titration Curves

2.) Example
(c) After the Equivalence Point ( 51.0 mL of EDTA)
Calculate [MgY2-]:
Original [Mn+] Original volume of
Mn+ solution
Original volume
solution
Volume titrant
added
Dilution effect
Moles Mg+ ≡ moles MgY2-
Only Difference

EDTA Titrations
EDTA Titration Curves

2.) Example
(c) After the Equivalence Point ( 51.0 mL of EDTA)
[Mg2+-] is given by the equilibrium expression using [EDTA] and [MgY2-]:

EDTA Titrations
EDTA Titration Curves

2.) Example
Final titration curve for 50.0 ml of 0.0500 M Mg2+ with 0.0500 m EDTA at pH 10.00.
Also shown is the titration of 50.0 mL of 0.0500 M Zn2+
Note: the equivalence point is sharper for Zn2+ vs. Mg2+. This is due to Zn2+ having a larger formation constant.
The completeness of these reactions is dependent on αY4- and correspondingly pH.
pH is an important factor in setting the completeness and selectivity of an EDTA titration

EDTA Titrations
Auxiliary Complexing Agents

1.) Metal Hydroxide
In general, as pH increases a titration of a metal ion with EDTA will have a higher Kf.
Larger change at the equivalence point.

Exception: If Mn+ reacts with OH- to form an insoluble metal hydroxide

Auxiliary Complexing Agents: a ligand can be added that complexes with Mn+ strong enough to prevent hydroxide formation.
Ammonia, tartrate, citrate or triethanolamine
Binds metal weaker than EDTA
Fraction of free metal ion (αM) depends on the equilibrium constants (β) or cumulative formation constants:
Use a new conditional formation constant that incorporates the fraction of free metal:

EDTA Titrations
Auxiliary Complexing Agents

2.) Illustration:
Titration of Cu+2 (CuSO4) with EDTA
Addition of Ammonia Buffer results in a dark blue solution
Cu(II)-ammonia complex is formed
Addition of EDTA displaces ammonia with corresponding color change
CuSO4
Cu-EDTA
Cu-ammonia

EDTA Titrations
Metal Ion Indicators

1.) Determination of EDTA Titration End Point
Four Methods:
Metal ion indicator
Mercury electrode
pH electrode
Ion-selective electrode

Metal Ion Indicator: a compound that changes color when it binds to a metal ion
Similar to pH indicator, which changes color with pH or as the compound binds H+

For an EDTA titration, the indicator must bind the metal ion less strongly than EDTA
Similar in concept to Auxiliary Complexing Agents
Needs to release metal ion to EDTA
Potential Measurements
(red)
(colorless)
(colorless)
(blue)
End Point indicated by a color change from red to blue

EDTA Titrations
Metal Ion Indicators

2.) Illustration
Titration of Mg2+ by EDTA
Eriochrome Black T Indicator
Addition of EDTA
Before Near After
Equivalence point

EDTA Titrations
Metal Ion Indicators

3.) Common Metal Ion Indicators
Most are pH indicators and can only be used over a given pH range

EDTA Titrations
Metal Ion Indicators

3.) Common Metal Ion Indicators
Useful pH ranges

EDTA Titrations
EDTA Titration Techniques

1.) Almost all elements can be determined by EDTA titration
Needs to be present at sufficient concentrations

Extensive Literature where techniques are listed in:
G. Schwarzenbach and H. Flaschka, “Complexometric Titrations”, Methuen:London, 1969.
H.A. Flaschka, “EDTA Titrations”, Pergamon Press:New York, 1959
C.N. Reilley, A.J. Bernard, Jr., and R. Puschel, In: L. Meites (ed.) “Handbook of Analytical Chemistry”, McGraw-Hill:New York, 1963; pp. 3-76 to 3-234.

Some Common Techniques used in these titrations include:
Direct Titrations
Back Titrations
Displacement Titrations
Indirect Titrations
Masking Agents

EDTA Titrations
EDTA Titration Techniques

2.) Direct Titrations
Analyte is buffered to appropriate pH and is titrated directly with EDTA

An auxiliary complexing agent may be required to prevent precipitation of metal hydroxide.

3.) Back Titrations
A known excess of EDTA is added to analyte
Free EDTA left over after all metal ion is bound with EDTA

The remaining excess of EDTA is then titrated with a standard solution of a second metal ion

Approach necessary if analyte:
precipitates in the presence of EDTA
Reacts slowly with EDTA
Blocks the indicator

Second metal ion must not displace analyte from EDTA

EDTA Titrations
EDTA Titration Techniques

4.) Displacement Titration
Used for some analytes that don’t have satisfactory metal ion indicators

Analyte (Mn+) is treated with excess Mg(EDTA)2-, causes release of Mg2+.

Amount of Mg2+ released is then determined by titration with a standard EDTA solution
Concentration of released Mg2+ equals [Mn+]

Requires:

EDTA Titrations
EDTA Titration Techniques

5.) Indirect Titration
Used to determine anions that precipitate with metal ions

Anion is precipitated from solution by addition of excess metal ion
ex. SO42- + excess Ba2+
Precipitate is filtered & washed

Precipitate is then reacted with excess EDTA to bring the metal ion back into solution

The excess EDTA is titrated with Mg2+ solution

[Total EDTA] = [MYn-4] + [Y4-] complex
free
Known
Titrate
determine

EDTA Titrations
EDTA Titration Techniques

6.) Masking Agents
A reagent added to prevent reaction of some metal ion with EDTA

Demasking: refers to the release of a metal ion from a masking agent
Al3+ is not available to bind EDTA because of the complex with F-
Requires:

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