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Using Iron Porphyrins as Models for Hemoglobin

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

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Using Iron Porphyrins as Models for Hemoglobin

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  1. Using Iron Porphyrins as Models for Hemoglobin The system:

  2. Key Features of Hemes • Fe oxidation state • Fe spin state • porphyrin oxidation state • porphyrin • hydrophobicity

  3. 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

  4. Sample for Evans’ Magnetic Susceptibility Method NMR tube Inside capillary: sample in CHCl3, 1) with imidazole 2) without imidazole Outside capillary: CDCl3 and CHCl3

  5. NMR Spectrum from Evans’ Method Inside capillary: sample in CHCl3, produces broad singlet for paramagnetically shifted CHCl3 below 7.3 ppm Outside capillary: CDCl3 and CHCl3 produces usual sharp singlet for 0.5% CHCl3 at 7.3 ppm 

  6. 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

  7. Shift of signal, in Hz mass susceptibility of solvent -a diamagnetic contribution, a (-) value Mass susceptibility (+) Magnetic field (400 MHz, or 400 x 106 Hz) Concentration of sample, in g/mL How does shift, , relate to a magnetization of paramagnetic sample? g = 3  0 c

  8. 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

  9. 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 corr1/2 = 2.828 (T corr) 1/2 N 2 eff = (n(n+2))1/2

  10. 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

  11. The Role of Axial Ligation and the Allosteric Effect in Hemoglobin O2 Binding

  12. 3d orbitals on Fe Spin State of Fe affects size of ion

  13. 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

  14. How Magnetic Nuclei Benefit NMR Experiments

  15. Use of Cr(acac)3as a Paramagnetic Relaxation Agent

  16. Use of Cr(acac)3as a Paramagnetic Relaxation Agent in 13C NMR With Cr(acac)3 (note: does not affect chemical shifts) With d1=6.0s (d1: relaxation time)

  17. Use of Paramagnetic NMR in Bioinorganic Systems v m One big mess of piled up H’s on protein!! v m m p p

  18. NMR Paramagnetic Shift Reagents Ground state electron configuration: [Xe] 4f7 6s2 Term Symbol: 8S7/2 how many unpaired e-? EuFOD :also called Eu(fod)3. Eu(OCC(CH3)3CHCOC3F7)3

  19. NMR Paramagnetic Shift Reagents: Eu vs Pr Using Eu(fod)3 oooh! Lovely!! With NO Shift rgt  Hmmm, not so pretty Huh? – signals shifted upfield with Pr Using Pr(fod)3

  20. MRI Contrast agents: same principles, applied to medicine • MRI Contrast Agents: observes differential magnetization of protons in different types of molecules that predominate in different tissues. The different magnetization signal intensities produce the contrast between tissues. • The nuclear magnetization is produced by the pulse sequence applied, by the density of nuclear spins sub-fractions (water vs fat protons) and by the spin-lattice relaxation time T1 and phase relaxation time T2 in each nuclear spin sub-fraction. T1 and T2 depend on tissues type. • MRI Contrast Agents interact with one sub-fraction type (usually that easily exchangeable protons, like water) to increase the T1 spin-lattice relaxation times. • The most commonly used compounds for contrast enhancement are gadolinium-based. • MRI contrast agents are used as oral or intravenous administration. Gadoteric acid Effect of contrast agent on images: Defect of the blood–brain barrier after stroke shown in MRI. T1-weighted images, left image without, right image with contrast medium administration.

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