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Pure manganese(III) 5,10,15,20-tetrakis(4-benzoic acid)porphyrin (MnTBAP) is not a superoxide dismutase mimic in aqueous systems: a case of structure–activity relationship as a watchdog mechanism in experimental therapeutics and biology

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Abstract

Superoxide is involved in a plethora of pathological and physiological processes via oxidative stress and/or signal transduction pathways. Superoxide dismutase (SOD) mimics have, thus, been actively sought for clinical and mechanistic purposes. Manganese(III) 5,10,15,20-tetrakis(4-benzoic acid)porphyrin (MnTBAP) is one of the most intensely explored “SOD mimics” in biology and medicine. However, we show here that this claimed SOD activity of MnTBAP in aqueous media is not corroborated by comprehensive structure–activity relationship studies for a wide set of Mn porphyrins and that MnTBAP from usual commercial sources contains different amounts of noninnocent trace impurities (Mn clusters), which inhibited xanthine oxidase and had SOD activity in their own right. In addition, the preparation and thorough characterization of a high-purity MnTBAP is presented for the first time and confirmed that pure MnTBAP has no SOD activity in aqueous medium. These findings call for an assessment of the relevance and suitability of using MnTBAP (or its impurities) as a mechanistic probe and antioxidant therapeutic; conclusions on the physiological and pathological role of superoxide derived from studies using MnTBAP of uncertain purity should be examined judiciously. An unequivocal distinction between the biological effects due to MnTBAP and that of its impurities can only be unambiguously made if a pure sample is/was used. This work also illustrates the contribution of fundamental structure–activity relationship studies not only for drug design and optimization, but also as a “watchdog” mechanism for checking/spotting eventual incongruence of drug activity in chemical and biological settings.

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Notes

  1. At pH 7.8 the carboxylato groups of MnTBAP are deprotonated and the resulting species bears an overall −3 or −4 charge depending on the oxidation state of the Mn: [MnIIITBAP]3− (six negative charges on the ring and three positive charges on the MnIII center) or [MnIITBAP]4− (six negative charges on the ring and two positive charges on the MnII center).

Abbreviations

DMF:

N,N-Dimethylformamide

EDTA:

Sodium salt of ethylenediaminetetraacetic acid

ESI:

Electrospray ionization

Fc:

Ferrocene

H2TBAP:

5,10,15,20-Tetrakis(4-benzoic acid)porphyrin

H2TBAP-TME:

5,10,15,20-Tetrakis(4-benzoic acid)porphyrin tetramethyl ester

IC20 :

Concentration that causes 20% inhibition of xanthine oxidase activity

MnTBAP:

Manganese(III) 5,10,15,20-tetrakis(4-benzoic acid)porphyrin

Mn(TBAP-TME)Cl:

Manganese(III) 5,10,15,20-tetrakis(4-benzoic acid)porphyrin tetramethyl ester chloride

MnTE-2-PyP5+ :

Manganese(III) 5,10,15,20-tetrakis(N-ethylpyridinium-2-yl)porphyrin

MnTnHex-2-PyP5+ :

Manganese(III) 5,10,15,20-tetrakis(N-n-hexylpyridinium-2-yl)porphyrin

MS:

Mass spectrometry

MS/MS:

Tandem mass spectrometry

NHE:

Normal hydrogen electrode

NMR:

Nuclear magnetic resonance

RNS:

Reactive nitrogen species

ROS:

Reactive oxygen species

SAR:

Structure–activity relationship

SOD:

Superoxide dismutase

X:

Xanthine

XO:

Xanthine oxidase

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Acknowledgements

The authors are grateful for the valuable discussions with Irwin Fridovich and Peter Hambright. This work was supported by the National Institutes of Health grants NIH IR21-ESO/3682, U19A167798-01, and NIH/NCI DCCC Core Grant 5-P30-CA14236-29.

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Authors and Affiliations

  1. Department of Radiation Oncology, Cancer Biology, Duke University Medical School, Research Drive, 201 MSRB I, Box 3455, Durham, NC, 27710, USA

    Júlio S. Rebouças & Ines Batinić-Haberle

  2. Department of Medicine, Duke University Medical School, Trent Drive, 5320 Hospital South, Box 3962, Durham, NC, 27710, USA

    Ivan Spasojević

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  1. Júlio S. Rebouças
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  2. Ivan Spasojević
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  3. Ines Batinić-Haberle
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Correspondence to Ines Batinić-Haberle.

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Rebouças, J.S., Spasojević, I. & Batinić-Haberle, I. Pure manganese(III) 5,10,15,20-tetrakis(4-benzoic acid)porphyrin (MnTBAP) is not a superoxide dismutase mimic in aqueous systems: a case of structure–activity relationship as a watchdog mechanism in experimental therapeutics and biology. J Biol Inorg Chem 13, 289–302 (2008). https://doi.org/10.1007/s00775-007-0324-9

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