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Covalent adduct formation between the plasmalogen-derived modification product 2-chlorohexadecanal and phloretin.

Üllen A, Nusshold C, Glasnov T, Saf R, Cantillo D, Eibinger G, Reicher H, Fauler G, Bernhart E, Hallstrom S, Kogelnik N, Zangger K, Oliver Kappe C, Malle E, Sattler W - Biochem. Pharmacol. (2015)

Bottom Line: Adduct characterization by high-resolution mass spectroscopy confirmed these results.In contrast to 2-ClHDA, the covalent 2-ClHDA-phloretin adduct was without adverse effects on MTT reduction (an indicator for metabolic activity), cellular adenine nucleotide content, and barrier function of brain microvascular endothelial cells (BMVEC).We here identify a detoxification pathway for a prototypic chlorinated fatty aldehyde (generated via the MPO axis) that compromises BBB function in vitro and in vivo.

View Article: PubMed Central - PubMed

Affiliation: Institute of Molecular Biology and Biochemistry, Medical University of Graz, Graz, Austria.

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Analyte recovery during 2-ClHDA-phloretin adduct formation. (A) Phloretin structure. The A- and B-ring is indicated. (B) DPPC liposomes containing 2-ClHDA and phloretin were incubated at 37 °C in PBS. (C) 2-ClHDA and phloretin were incubated in acetonitrile (containing 0.1% triethylamine) at 37 °C. (D) 2-ClHDA and phloretin were incubated in PBS containing 250 μg HDL protein/ml at 37 °C. At the indicated time points aliquots of the reaction mixture were extracted, and either converted to the corresponding PFB-oxime derivatives and analyzed by NICI–GC–MS using 2-Cl[13C8]HDA as internal standard (B) or were quantitated by HPLC analysis (C and D) using external calibration. Results are presented as percentage recovery of the corresponding analytes. Results represent mean ± SD from triplicate experiments.
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fig0005: Analyte recovery during 2-ClHDA-phloretin adduct formation. (A) Phloretin structure. The A- and B-ring is indicated. (B) DPPC liposomes containing 2-ClHDA and phloretin were incubated at 37 °C in PBS. (C) 2-ClHDA and phloretin were incubated in acetonitrile (containing 0.1% triethylamine) at 37 °C. (D) 2-ClHDA and phloretin were incubated in PBS containing 250 μg HDL protein/ml at 37 °C. At the indicated time points aliquots of the reaction mixture were extracted, and either converted to the corresponding PFB-oxime derivatives and analyzed by NICI–GC–MS using 2-Cl[13C8]HDA as internal standard (B) or were quantitated by HPLC analysis (C and D) using external calibration. Results are presented as percentage recovery of the corresponding analytes. Results represent mean ± SD from triplicate experiments.

Mentions: To monitor the rate of 2-ClHDA scavenging by phloretin (structure shown in Fig. 1A) the reactants were incorporated in DPPC liposomes at equimolar ratios. At the indicated times aliquots of the reaction mixture were removed, extracted, derivatized, and analyzed by NICI–GC–MS. These analyses revealed that 2-ClHDA concentrations decreased time-dependently (Fig. 1B) which could be fitted according to a first order decay (Ct = Co × e−kt; r2 = 0.98). Of note, the constants reported here are observed rate constants, not second order rate constants. Under the experimental conditions used the half-life (τ/2) of 2-ClHDA was 120 min, with a calculated decay rate of 5.9 × 10−3 min−1. At the latest time point (24 h) virtually all 2-ClHDA was consumed.


Covalent adduct formation between the plasmalogen-derived modification product 2-chlorohexadecanal and phloretin.

Üllen A, Nusshold C, Glasnov T, Saf R, Cantillo D, Eibinger G, Reicher H, Fauler G, Bernhart E, Hallstrom S, Kogelnik N, Zangger K, Oliver Kappe C, Malle E, Sattler W - Biochem. Pharmacol. (2015)

Analyte recovery during 2-ClHDA-phloretin adduct formation. (A) Phloretin structure. The A- and B-ring is indicated. (B) DPPC liposomes containing 2-ClHDA and phloretin were incubated at 37 °C in PBS. (C) 2-ClHDA and phloretin were incubated in acetonitrile (containing 0.1% triethylamine) at 37 °C. (D) 2-ClHDA and phloretin were incubated in PBS containing 250 μg HDL protein/ml at 37 °C. At the indicated time points aliquots of the reaction mixture were extracted, and either converted to the corresponding PFB-oxime derivatives and analyzed by NICI–GC–MS using 2-Cl[13C8]HDA as internal standard (B) or were quantitated by HPLC analysis (C and D) using external calibration. Results are presented as percentage recovery of the corresponding analytes. Results represent mean ± SD from triplicate experiments.
© Copyright Policy - CC BY
Related In: Results  -  Collection

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Show All Figures
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fig0005: Analyte recovery during 2-ClHDA-phloretin adduct formation. (A) Phloretin structure. The A- and B-ring is indicated. (B) DPPC liposomes containing 2-ClHDA and phloretin were incubated at 37 °C in PBS. (C) 2-ClHDA and phloretin were incubated in acetonitrile (containing 0.1% triethylamine) at 37 °C. (D) 2-ClHDA and phloretin were incubated in PBS containing 250 μg HDL protein/ml at 37 °C. At the indicated time points aliquots of the reaction mixture were extracted, and either converted to the corresponding PFB-oxime derivatives and analyzed by NICI–GC–MS using 2-Cl[13C8]HDA as internal standard (B) or were quantitated by HPLC analysis (C and D) using external calibration. Results are presented as percentage recovery of the corresponding analytes. Results represent mean ± SD from triplicate experiments.
Mentions: To monitor the rate of 2-ClHDA scavenging by phloretin (structure shown in Fig. 1A) the reactants were incorporated in DPPC liposomes at equimolar ratios. At the indicated times aliquots of the reaction mixture were removed, extracted, derivatized, and analyzed by NICI–GC–MS. These analyses revealed that 2-ClHDA concentrations decreased time-dependently (Fig. 1B) which could be fitted according to a first order decay (Ct = Co × e−kt; r2 = 0.98). Of note, the constants reported here are observed rate constants, not second order rate constants. Under the experimental conditions used the half-life (τ/2) of 2-ClHDA was 120 min, with a calculated decay rate of 5.9 × 10−3 min−1. At the latest time point (24 h) virtually all 2-ClHDA was consumed.

Bottom Line: Adduct characterization by high-resolution mass spectroscopy confirmed these results.In contrast to 2-ClHDA, the covalent 2-ClHDA-phloretin adduct was without adverse effects on MTT reduction (an indicator for metabolic activity), cellular adenine nucleotide content, and barrier function of brain microvascular endothelial cells (BMVEC).We here identify a detoxification pathway for a prototypic chlorinated fatty aldehyde (generated via the MPO axis) that compromises BBB function in vitro and in vivo.

View Article: PubMed Central - PubMed

Affiliation: Institute of Molecular Biology and Biochemistry, Medical University of Graz, Graz, Austria.

Show MeSH
Related in: MedlinePlus