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Structural isomerization of synephrine influences its uptake and ensuing glutathione depletion in rat-isolated cardiomyocytes

  • Toxicokinetics and Metabolism
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Abstract

Synephrine is a natural compound, frequently added to ephedra-free dietary supplements for weight-loss, due to its effects as a nonspecific adrenergic agonist. Though only p-synephrine has been documented in plants, the presence of m-synephrine has also been reported in weight-loss products. The use of synephrine in dietary supplements was accompanied by reports of adverse effects, especially at the cardiovascular level. It is well known that the imbalance in cardiac glutathione levels can increase the risk of cardiomyopathy. The present work aimed to study the role of organic cation-mediated transport of m- and p-synephrine and the possibility that p- and m-synephrine induce intracellular changes in glutathione levels in calcium-tolerant freshly isolated cardiomyocytes from adult rat. After a 3 h incubation with 1 mM p- or m-synephrine, the intracellular content of synephrine was measured by gas chromatography/ion trap-mass spectrometry (GC/IT-MS); cell viability and intracellular glutathione levels were also determined. To evaluate the potential protective effects of antioxidants against the adverse effects elicited by m-synephrine, cells were pre-incubated for 30 min with Tiron (100 μM) or N-acetyl-cysteine (NAC) (1 mM). To assess the influence of α1-adrenoceptors activation in glutathione depletion, a study with prazosin (100 nM) was also performed. The results obtained provide evidence that organic cation transporters OCT3 and OCT1 play a major role in m- and p-synephrine-mediated transport into the cardiomyocytes. The importance of these transporters seems similar for both isomers, although p-synephrine enters more into the cardiomyocytes. Furthermore, only m-synephrine induced intracellular total glutathione (GSHt) and reduced glutathione (GSH) depletion. NAC and Tiron were able to counteract the m-synephrine-induced GSH and GSHt decrease. On the other hand, the incubation with prazosin was not able to change m-synephrine-induced glutathione depletion showing that this effect is independent of α1-adrenoceptor stimulation. In conclusion, both positional isomers require OCT3 and OCT1-mediated transport to enter into the cardiomyocytes; however, the hydroxyl group in the p-position favours the OCT-mediated transport into cardiomyocytes. Furthermore, the structural isomerization of synephrine influences its toxicological profile since only m-synephrine caused GSH depletion.

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Abbreviations

CORT:

Corticosterone

DMSO:

Dimethyl sulphoxide

DTNB:

5,5-dithio-bis(2-nitrobenzoic acid)

FDA :

Food and Drug Administration

GC/IT-MS:

Gas chromatography/ion trap-mass spectrometry

GR:

Glutathione reductase

GSH:

Reduced glutathione

GSHt:

Total glutathione

GSSG:

Glutathione disulphide

HClO4 :

Perchloric acid

HEPES:

N-(2-hydroxyethyl) piperazine-N-(2-ethanesulphonic acid)

IS:

Internal standard

LDH:

Lactate dehydrogenase

MRP1:

Multidrug resistance protein 1

NAC:

N-acetyl-cysteine

NADPH:

Reduced nicotinamide adenine dinucleotide phosphate

OCT:

Organic cation transporter

OCTN:

Novel organic cation transporter

PMAT:

Plasma membrane monoamine transporter

QUIN:

Quinidine

ROS:

Reactive oxygen species

SD:

Standard deviation

SPE:

Solid-phase extraction

TFAA:

Trifluoroacetic anhydride

Tiron:

4,5-dihydroxy-1,3-benzene disulphonic acid

β-NADH:

Reduced β-nicotinamide adenine dinucleotide

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Acknowledgments

This work received financial support from the Portuguese State through “Fundação para a Ciência e Tecnologia” (FCT) (project PPCDT/SAU-OBS/55849/2004). LGR thanks FCT for her PhD grant (SFRH/BD/63473/2009) and VMC thanks FCT for her Post-PhD grant (SFRH/BPD/63746/2009).

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Authors and Affiliations

  1. REQUIMTE, Laboratório de Toxicologia, Departamento de Ciências Biológicas, Faculdade de Farmácia, Universidade do Porto, Rua Aníbal Cunha, 164, 4099-030, Porto, Portugal

    Luciana Grazziotin Rossato, Vera Marisa Costa, Paula Guedes de Pinho, Félix Carvalho, Maria de Lourdes Bastos & Fernando Remião

  2. REQUIMTE, Departamento de Toxicologia, Faculdade de Farmácia, Universidade do Porto, Rua Aníbal Cunha, 164, 4099-030, Porto, Portugal

    Luciana Grazziotin Rossato & Fernando Remião

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  1. Luciana Grazziotin Rossato
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  2. Vera Marisa Costa
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  3. Paula Guedes de Pinho
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  4. Félix Carvalho
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  5. Maria de Lourdes Bastos
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  6. Fernando Remião
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Correspondence to Luciana Grazziotin Rossato or Fernando Remião.

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Rossato, L.G., Costa, V.M., de Pinho, P.G. et al. Structural isomerization of synephrine influences its uptake and ensuing glutathione depletion in rat-isolated cardiomyocytes. Arch Toxicol 85, 929–939 (2011). https://doi.org/10.1007/s00204-010-0630-9

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  • DOI: https://doi.org/10.1007/s00204-010-0630-9

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