Analysis Of Porphyrins And Metalloporphyrins

It is often important to know the concentration in solution of a particular porphyrin or metalloporphyrin. Since this class of molecules often contains various kinds and amounts of nonporphyrin material that are isolated along with the porphyrin itself, simply weighing out a given amount of compound may give inaccurate results. With water-soluble porphyrins, water molecules are usually indicated by chemical analysis of the isolated solids. The anionic water-soluble H2-TPPS4 (Figure 1) analyzes as Na4H2-TPPS4-10 H2O when oven dried at 110°C (83), the cationic pyri-dinium compound as H2-TMPyP(4)Cl4-4 H2O, various phenyl/(4-sulfonatophenyl), and phenyl/(N-methyl-4-pyridyl) por-phyrins contain from 2 to 11 moles of water, often in nonstoichiometric amounts, as Zn-TPPS4 analyzes for 16.6 H2O. Thermo-gravimetric work (53) indicated that the latter compound gradually lost weight up to 200°C, a constant-weight region was found in the 220° to 400°C range, while rapid decomposition was noted above 400°C. Compounds prepared at different times sometimes contain differing amounts of water. The Fe(III)-TMPyP(4) is a monomer in acid, but when precipitated from acid solution by the addition of 1 M HClO4, Mossbauer spectra on the solid indicated 85%

monomer and 15% of the ^-oxo-bridged P-Fe-O-FeP dimer (91). The CHN analyses and corresponding ratios are always helpful, but the moles of water are usually determined by calculation based on the observed values. Crystal structures of water-insoluble porphyrins usually contain several molecules of the crystallizing solvent, and tetra-arylporphyrin crystals have channels throughout the solid that occlude solvent molecules and act as "porphyrin sponges" (19). On new compounds, most authors report CHN and metal analyses, the absorption spectra and extinction coefficients, infrared (IR) band positions, the analyzed 1H nuclear magnetic resonance (NMR) data, and the major peaks obtained by mass spectrometry.

It is better to determine solution concentrations based on known extinction coefficients, than to rely on calculations based on the weight of the sample. Due to aggregation effects, the spectra should be measured at concentration levels similar to the value quoted in the literature. DiNello and Chang (29) give band positions and extinction coefficients in CHCl3 for the free bases of modified natural porphyrin derivatives, and for the corresponding Fe(II) pyridine hemochromes, measured in 4 M pyridine-0.2 M KOH with the addition of small amounts of the reducing agent sodium dithionite. Caughey and coworkers (23) have spectra on other free base 3,8-disubstituted deuteroporphyrin DMEs and metallo derivatives (22). Caughey et al. have compiled data on isomers of uroporphyrins and copropor-phyrins and other biologically important molecules (21). Fuhrhop and Smith (44) have five tables of extinction coefficients of various porphyrins and metalloporphyrins in organic solvents and in aqueous sodium dodecyl sulfate solutions. The spectra of centrally N-alkylated porphyrins and met-alloporphyrins are found in Lavallee's monograph (71), and James and coworkers

(83) give data on well-characterized water-soluble porphyrins and their precursors. Methods for the determination of por-phyrins in biological material have been reviewed (103).

Finally, there are a number of laboratory procedures to determine porphyrin concentrations. In Drabkin's method, iron protoporphyrin IX is digested with hydrogen peroxide in basic solution, the liberat

Porphyrin Mass Spectrometry

Figure 1. The structures of some water-soluble porphyrins and several of their precursors. Each compound is the porphyrin with the indicated substituents on the four meso (5, 10, 15 and 20) positions. H2-TPP: meso-tetraphenylporphyrin; H2-TPyP(4), meso-tetrakis(4-pyridyl)porphyrin; H2-TMPyP(X): the ortho (2), meta (3) and para (4) isomers of meso-tetrakis(N-methyl-X-pyridyl)porphyrin; H2-TPPC4, meso-tetrakis(4-carboxyphenyl)porphyrin; H2-TPPS4: meso-tetrakis(4-sul-fonatophenyl)porphyrin; H2-TAPP, meso-tetrakis(4-N,N,N-trimethylanilinium)porphyrin. A fuller explanation of the nomenclature of porphyrins can be found in Chapter 2.

Figure 1. The structures of some water-soluble porphyrins and several of their precursors. Each compound is the porphyrin with the indicated substituents on the four meso (5, 10, 15 and 20) positions. H2-TPP: meso-tetraphenylporphyrin; H2-TPyP(4), meso-tetrakis(4-pyridyl)porphyrin; H2-TMPyP(X): the ortho (2), meta (3) and para (4) isomers of meso-tetrakis(N-methyl-X-pyridyl)porphyrin; H2-TPPC4, meso-tetrakis(4-carboxyphenyl)porphyrin; H2-TPPS4: meso-tetrakis(4-sul-fonatophenyl)porphyrin; H2-TAPP, meso-tetrakis(4-N,N,N-trimethylanilinium)porphyrin. A fuller explanation of the nomenclature of porphyrins can be found in Chapter 2.

ed ferric iron reduced with ascorbate, and the Fe(II) spectrophotometrically analyzed with ¿»-phenanthroline (32). With metallo-TPPS4 complexes, approximately 10-mg samples were digested with a 3:1:1 mixture of H2SO4-HNO3-HClO4, and the metal then determined by titration with EDTA (53). Brisbin and coworkers (13) spectro-photometrically determined the concentration of protoporphyrin IX DME to ± 2%. To a constant amount of porphyrin (approximately 10'5 M) in acetic acid, higher and lower concentrations of standardized metal acetate solutions (divalent Zn, Co, Ni, Fe, and Cu) were added. After heating to completely form the 1:1 complex, appropriate plots of absorbance versus metal ion concentration indicate the point at which the concentration of the metal is equal to the concentration of the porphyrin. This method can also be applied to determine the concentration of the metals (2—5 x 10-5 M) by titration with a known concentration of porphyrin. Petro and Marzilli (93) determined the molar extinction coefficients of a series of cation-ic porphyrins by adding excess standardized Zn(II) to aqueous solutions of the por-phyrins at pH 12.0. After the reaction was complete, the pH was brought to 9.0, and excess zinc was determined with the colori-metric zinc reagent, zincon. The precision of this method was approximately 3%.

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