True or false?
Cyanide is poisonous because it binds more tightly
to the iron in hemoglobin than does O2 and cause suffocation.
The Singapore government gets it right:
“Blood Agents: Cyanide-containing compound, affect
body functions by poisoning the enzymes, Cytochrome
Oxidise. Hence preventing the normal utilization of
oxygen by the cells and causing rapid damage to body
tissues.”
About Chemical Warfare Agents
False
Let’s do the calculation.
•You have ~ 2g Fe in your body; about 1.5g of Fe is in your blood as hemoglobin.
•1.5 g Fe = 0.03 mol.
•Toxicity of cyanide:
The LD50 for ingestion is 50-200 milligrams, or 1-3 milligrams per kilogram of body
weight, calculated as hydrogen cyanide. The LC50 for gaseous hydrogen cyanide is 100-300 parts per million.
Inhalation of cyanide in this range results in death within 10-60 minutes, with death coming more quickly as the
concentration increases. Inhalation of 2,000 parts per million hydrogen cyanide causes death within one minute.
•So, if you weigh 60 kg, between 60-180 mg could be a toxic dose.
•Or, 0.06-0.18g = 0.002-0.007 mol CN- (as HCN) vs 0.03 mol Fe
•That amount of cyanide would block only ~10% of O2 binding sites in
hemoglobin, not enough to kill you.
The toxicity of Cyanide is because of its strong binding to Fe in heme in
other critical Heme-proteins.
The Many Role of Hemes
• oxygen carrier (hemoglobin)
• electron transfer (cytochromes a,b,c, etc, in respiratory chain)
• cytochrome oxidase (mitochondrial electron transport chain,
oxygen is terminal electron acceptor and is reduced to water)
• detoxification (cytochrome P450, catalase)
• hydroxylation (cytochrome P450 in hormone production)
Complex III
matrix
Q
HEME 1
2Q
QH2
HEME 2
2 x QH2
cytosol
c1 HEME 3
c
HEME 4
Cyt c
Complex IV – Cytochrome C Oxidase
Fe(III) Fe(II)
HEME 1
Cu(II)—Cu(I)  Cu(I)—Cu(I)
HEME 2
HEME 3
HEME 4
Cu(II)  Cu(I)
Using Iron Porphyrins as Models for Hemoglobin
The system:
Key Features of Hemes
•
•
•
•
Fe oxidation state
Fe spin state
porphyrin oxidation state
porphyrin
hydrophobicity
How will the spin state of Fe(porphyrin) complexes change on binding imidazole?
Intermediate Spin
S = 3/2
n=3
High Spin
S = 5/2
n=5
Low Spin
S = 1/2
n=1
Sample for Evans’ Magnetic Susceptibility Method
NMR tube
Inside capillary: sample in CHCl3,
1) with imidazole
2) without imidazole
Outside capillary: 99.5 %D CDCl3
NMR Spectrum from Evans’ Method
Inside capillary: sample in CHCl3,
produces broad singlet for
paramagnetically shifted CHCl3
below 7.3 ppm
Outside capillary: 99.5 %D CDCl3
produces usual sharp singlet for
0.5% CHCl3 at 7.3 ppm

Why is H resonance in CHCl3 shifted downfield and broadened?
• pseudocontact and contact terms
• addition of new small magnetic field to local magnetic fields
of neighboring nuclei
is used in NMR Shift Reagents to “de-tangle” complicated spectra
How does shift, , relate to a magnetization of paramagnetic sample?
Shift of signal, in Hz
g = 3  0
 c
Mass susceptibility (+)
mass susceptibility of
solvent
-a diamagnetic contribution,
a (-) value
Magnetic field
(400 MHz, or 400 x 106 Hz)
Concentration of sample,
in g/mL
Magnetic field lines of flux
Magnetic field lines
affected by a paramagnetic
substance: attracts
Susceptibility, X > 0
Magnetic field lines
affected by a diamagnetic
substance: repels
Susceptibility, X < 0
How does mass susceptibility, g , relate to unpaired electrons in a
paramagnetic sample?
Mass susceptibility
g x (Mol. Wt.)
= M
Molar susceptibility
corr = M - diamagnetic corrections
where diamagnetic corrections for Fe, porphyrin, Cl, imidazole,
a negative number!
eff = 3 R T corr
N 2
eff = (n(n+2))1/2
1/2
= 2.828 (T corr ) 1/2
Diamagnetic Corrections (cgs units)
Xo (CHCl3) = - 4.97 x 10-7 cgs
Porphyrin:
TPP= -700 x 10-6 cgs
TTP= -753 x 10-6 cgs
TClPP= -760 x 10-6 cgs
Fe = -13 x 10-6 cgs
Cl = -20 x 10-6 cgs
Imidazole = -38 x 10-6 cgs
Characterization by Cyclic Voltammetry
1. How will the Ered of Fe(porphyrin) complexes vary
with the porphyrin?
2. How will the Ered of Fe(porphyrin) complexes change
on binding imidazole?
3. Will the Ered potentials also reflect a change in spin
state?
Intermediate Spin
S = 3/2
n=3
High Spin
S = 5/2
n=5
Low Spin
S = 1/2
n=1
The Role of Axial Ligation and the Allosteric Effect in Hemoglobin O2 Binding
3d orbitals
on Fe
Spin State of Fe affects size of ion
Large, high spin
Fe(2+):
In T state,
transmitted by His
on protein helix
Small, low spin
Fe(2+):
In R state,
transmitted to His
on protein helix