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Nitric Oxide Production from Synthesized Ruthenium (III) Porphyrins
Matthew J. Ranaghan,
Jeannette Suh, Stephen K. O’Shea, and Cliff J. Timpson
Roger Williams University, One Old Ferry Road, Bristol, RI 02809
Abstract
Discussion
One of the regulatory processes of nitrite in mammalian systems is by its reaction with iron found in porphyrin
systems. One important reaction is the production of an Iron (II) nitrosyl from Iron (III) and a nitrite ion. This
research investigated Ru (III) porphyrin systems, as it is a possible candidate in blood substitutes and in therapeutic
drugs as cofactors. The Ru (III) porphyrin base was varied between octaethylporphyrin, tetraphenylporphyrin, and
saturated ring cyclam (1,4,8,11-tetraazacyclotetradecane). The addition of nitrite to varying Ru (III) macrocycles will
be monitored with repetitive UV-vis spectroscopy scans in order to determine the rate of reaction for the formation
of Ru(II) NO macrocycles. The varyation of the E1/2 redox potential of the Ru (III) center is to be investigated by
cyclic voltammetry and correlated to the reaction rate to elucidate the mechanism.
Research of Ru porphyrins and their production of nitric oxide was investigated in order to elucidate whether the mechanism is similar to that
of Fe in living organisms. Normal oxidation states for Fe are +2/+3 (unbound/bound Oxygen respectively). The dependence of NO binding to
the metal center does not differ only by the metal of interest, but also relies on the exterior ligands10 of the porphyrin system. Therefore, the
macrocyclic structure was varied between octaethylporphyrin (OEP), tetraphenylporphyrin (TPP) and saturated ring cyclam (1,4,8,11tetraazacyclotetradecane). The kinetics of NO2- binding to the Ruthenium were studied by varying the exterior porphyrin ligands, and slight
variation in the ring structure (cyclam). The results show that nitric oxide production is possible through the presence of nitrite and a
triphenylphosphine (PPh3) substrate that removes an oxygen from the activated nitrite Ruthenium complex center of the porphyrin. The
Ruthenium metal is also reduced upon the formation of the nitrosyl.
The proposed mechanism is reported by a correlation of the authentic Ru(II)NO macrocycle and the final spectra of the monitored reaction.
Triphenylphosphine oxide (OPPh3) was qualitatively observed by gas chromatographic methods. The mechanism of nitrite coordination and
oxidation of the PPh3 to OPPh3 was shown to be redox dependant. The activated complex undergoes reduction from RuIII to RuII corellating
with the rate of nitrosyl formation.
Introduction
Rate (s-1)
1.09 x 10-5
1.02 x 10-5
0.94 x 10-5
Complex
Ru(TPP)a
Ru(OEP)b
Ru(cyclam)b
Porphyrins play a vital role in the mechanism of life. A class of porphyrins (or hemes) that is utilized by multicellular
organisms is found in hemoglobin and myoglobin. Both of these are used for oxygen (O2) and carbon dioxide
(CO2) transportation throughout the body. Cytochromes are a class of monomeric active porphyrin systems
involved in metabolic redox pathways, and are responsible for the generation of nitric oxide (NO) in the body. The
NO molecule is a highly reactive free radical that is used for specific purposes in biological systems, such as in
immune system response mechanisms, the cardiovascular and nervous system1, as well as messenger signals in
and between cells.2
iron ion is found at the center of heme structures of living organisms. Ru (II) heme analogs are found to be more
stable than their Fe (II) counterparts, while behaving with similar chemistry to the Fe derivatives since Ru is a
chemically similar metal.3 The importance of investigating a Ru metal center in place of an Fe center is because of
the interest in whether the Ru complexes will be stable enough to be used for blood substitutes.4 Stripping away
the protein shell of Cytochromes or Hemoglobin will leave just the active metal porphyrin center.5 Observing the
subtle effects at this site will give a more intuitive and clearer perspective of the mechanism of NO generation.
Here, we propose another possible source of NO that originates from nitrite (NO2-) interacting with the metal center
of hemes. Once attached to the metal center, an oxygen atom is transferred from the molecule to a substrate
molecule. Investigations of the interaction between an Fe active site with nitrite and the formation of a nitrosyl
ligand have been thoroughly researched.6 Therefore, research was focused on the investigation of the interaction
of Nitrite with the Ru (II) porphyrin chloride.
a – in 0.1M TBAH/CH2Cl2
E1/2 RuII/III (V)
0.21 (vs. SSCE)
0.08 (vs. SSCE)
-0.15 (vs. NHE)
b - in 0.1M TBAH/CH3CN
Alternate Ru(TPP)(NO) Generation
Complex Rate vs. E1/2
3
0.3
Pseudo First Order Kinetics of
Alternate Ru(TPP)(NO)
5
4.5
LN (Ao - A¥)/(At - A¥)
2
4
y = 0.094x - 1.7328
R2 = 0.8844
3.5
3
0.2
2.5
2
1.5
y = 241124x - 2.4048
2
R = 0.9855
1
0.5
0.1
0
0
10
1.5
20
30
40
50
60
70
E1/2 (V)
Absorbance (Au)
2.5
Time (nm)
0
9.00E-06
1
9.50E-06
1.00E-05
1.05E-05
1.10E-05
0.5
-0.1
0
450
500
550
600
650
Wavelength (nm)
700
-0.2
Rate (s-1)
[Ru(cyclam)NO] Generation
2.5
Pseudo First Order Kinetics of [Ru(cyclam)NO]
1.4
LN (Ao - A¥ )/(At - A¥ )
1.5
Conclusions
1.2
y = 0.0098x - 0.1384
R2 = 0.9473
1
0.8
0.6
0.4
0.2
0
0
20
40
60
80
100
120
140
Time (min)
1
0.5
0
400
420
440
460
480
500
520
540
560
580
600
Wavelength (nm)
0.8
Pseudo First Order Kinetics of Ru(OEP)NO
1.8
0.7
LN (Ao - A¥ )/(At - A¥ )
1.6
0.6
0.5
1.4
1.2
1
0.8
y = 0.0102x + 0.1819
2
R = 0.9745
0.6
0.4
0.2
0
0
20
40
60
80
100
120
140
160
Time (min)
0.4
0.3
0.2
0.1
500
520
540
560
580
• Ruthenium porphyrins are able to bind and produce Nitric Oxide through the aid of a substrate.
• Nitric Oxide is produced by the reduction of an oxygen molecule to a triphenylphosphine substrate.
• Exterior porphyrin ligands are important for increasing or decreasing the overall affinity of Nitrite to the Ruthenium center and the kinetics
of the reaction.
• Reasonable mechanism since nitrite is heavily present in the environment.
•The proposed mechanism of nitrite coordination and the oxidation of PPh3 is shown to be qualitatively correct from the presence of OPPh3
and the relationships between the spectra.
References
Ru(OEP)NO Generation
Absorbance (Au)
600
620
Wavelength (nm)
Ru(TPP)NO Generation
2.5
Pseudo First Order Kinetics of Ru(TPP)NO
1.
2.
3.
4.
Lorkovic I.M.; Ford, P.C. Inorg Chem. 1998, 38, 1467-1473.
Stryer, L. 1995. Biochemistry (4th ed.). Freeman. pg. 732.
Miranda, K.M.; Bu, X.; Lorkovic, I.; Ford, P.C. Inorg. Chem. 1997, 36, 4838-4848.
(a) Information obtained in a private conversation with Dr. O’Shea (November 14, 2002) (b) Zampieri, R.C.; Von Poelhsitz, G.; Batista, A.A.;
Nascimento, O.R.; Ellena, J.; Castellano, E. E. J. Inorg. Biochem. 2002, 92, 82-88.
5. O’Shea, S.K.; Wang, W.; Wade, R.S.; Castro, C.E. J. Org. Chem. 1996, 61, 6388-6395.
6. Kadish, K.M.; Adamian V.A.; Van Caemelbecke E.; Tan, Z.; Tagliatesta, P.; Bianco, P.; Boschi, T.; Yi, G.B.; Khan, M.A. Richter-Addo, G.B.
Inorg. Chem. 1996, 35, 1343-1348.
7. Alder, A.D.; Longo, F.R.; Finarelli, J.D.; Goldmacher, J.; Assour, J.; Korsakoff, L. J. Org. Chem. 1967, 32, 472.
8. Ariel, S.; Dolphin, D.; Domazetis, G.; James, B. R.; Leung, T. W.; Rettig, S. J.; Trotter, J.; Williams, G. M. Can. J. Chem 1984, 62, 755-762.
9. Lang, D.R.; Davis, J. A.; Lopes, L.; Ferro, A.; Vasconcellos, L.; Franco, D.; Tfouni, E.; Weiraszko, A.; Clarke, M. Inorg. Chem. 2000, 39,
2294-2300.
10. Zavarine, I. S.; Kini, A.D.; Morimoto, B.H.; Kubiak, C.P. J. Phys. Chem 1998, 102, 7287-7297.
1.2
2
LN (Ao - A¥ )/(At - A¥ )
Spectroscopic grade solvents (Aldrich, Fisher) and reagents (Aldrich, Frontier Scientific) were obtained
commercially and used as supplied. The complex meso-Tetraphenylporphine (TPP) was prepared according to
the procedure previously reported by Adler.7 Purification of TPP was carried out on activated alumina (mesh 80 200) column chromatography with chloroform as the eluent. The Octaethylporphine (OEP) complex was prepared
according to the procedure described by Ariel.8 The cyclam complex was prepared according to the procedure
described by Lang.9 The Alternate TPP-NO complex was prepared in according to the synthesis described by
Kadish.6 UV-Vis spectra and kinetic data were collected on a Hewlett-Packard HP-8453 Diode Array
spectrophotometer and gas chromatographic data was collected on the Perkin Elmer Sigma 300 Capillary
Chromatograph.
Absorbance (Au)
Methods and Materials
Absorbance (Au)
2
1.5
1
y = 0.0109x - 0.0233
R2 = 0.9592
0.8
0.6
0.4
0.2
0
0
10
20
30
40
50
60
70
80
90
100
Time (min)
1
0.5
0
440
490
540
Wavelength (nm)
590
640
Acknowledgments
This work was supported by the Office of Academic Affairs Grants, the Undergraduate
Research Program at Roger Williams University, and the Rhode Island NIH Brin grant.
Thanks to Meghan Gordon and Christopher Mucci for their help.
www.rwu.edu