Characterization of a chemically reactive propranolol metabolite that binds to microsomal proteins of rat liver

Shizuo Narimatsu, Toshiyuki Watanabe, Yasuhiro Masubuchi, Toshiharu Horie, Yoshito Kumagai, Arthur K. Cho, Susumu Imaoka, Yoshihiko Funae, Tsutomu Ishikawa, Tokuji Suzuki

Research output: Contribution to journalArticle

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Abstract

We have characterized a chemically reactive propranolol (PL) metabolite which binds to proteins in rat liver microsomes. During incubation with rat liver microsomes (1 mg of protein) fortified with an NADPH-generating system, 4-hydroxypropranolol (4-OH-PL) quickly disappeared from the reaction medium, but none of the possible metabolite peaks was detected under the high-performance liquid chromatographic conditions used. The consumption of 4-OH-PL depended on microsomes and NADPH. The reaction was not affected by inhibitors of cytochrome P450 or FAD monooxygenase, but was markedly diminished in the presence of cytosol and ascorbic acid. The effect of cytosol was inhibited by potassium cyanide but not by sodium benzoate or dimethyl sulfoxide, and was also not affected by heating at 60°C for 30 min, suggesting that superoxide (SO) ion was involved in the reaction and that it was blocked by superoxide dismutase (SOD) present in the cytosol. Cu,Zn-SOD, purified from cytosol, effectively mimicked the suppressive effect of cytosol. Incubation of 4-OH-PL in an SO-generating system of xanthine and xanthine oxidase generated 1,4-naphthoquinone (1,4-NQ), which was identified by TLC, HPLC, and GC/MS. 1,4-NQ was also formed in microsomal incubates containing NADPH and small amounts of microsomes (below 0.1 mg of protein). These results indicate that 4-OH-PL is converted by SO, or some reactive oxygen species derived from it, to 1,4-NQ which binds to proteins and is one of the reactive metabolites of PL.

Original languageEnglish
Pages (from-to)721-728
Number of pages8
JournalChemical Research in Toxicology
Volume8
Issue number5
Publication statusPublished - 1995
Externally publishedYes

Fingerprint

Metabolites
Propranolol
Liver
Rats
Cytosol
NADP
Superoxides
Proteins
Liver Microsomes
Microsomes
dimethylaniline monooxygenase (N-oxide forming)
Sodium Benzoate
Potassium Cyanide
Xanthine
Xanthine Oxidase
Dimethyl Sulfoxide
Heating
Cytochrome P-450 Enzyme System
Ascorbic Acid
Superoxide Dismutase

ASJC Scopus subject areas

  • Drug Discovery
  • Organic Chemistry
  • Chemistry(all)
  • Toxicology
  • Health, Toxicology and Mutagenesis

Cite this

Narimatsu, S., Watanabe, T., Masubuchi, Y., Horie, T., Kumagai, Y., Cho, A. K., ... Suzuki, T. (1995). Characterization of a chemically reactive propranolol metabolite that binds to microsomal proteins of rat liver. Chemical Research in Toxicology, 8(5), 721-728.

Characterization of a chemically reactive propranolol metabolite that binds to microsomal proteins of rat liver. / Narimatsu, Shizuo; Watanabe, Toshiyuki; Masubuchi, Yasuhiro; Horie, Toshiharu; Kumagai, Yoshito; Cho, Arthur K.; Imaoka, Susumu; Funae, Yoshihiko; Ishikawa, Tsutomu; Suzuki, Tokuji.

In: Chemical Research in Toxicology, Vol. 8, No. 5, 1995, p. 721-728.

Research output: Contribution to journalArticle

Narimatsu, S, Watanabe, T, Masubuchi, Y, Horie, T, Kumagai, Y, Cho, AK, Imaoka, S, Funae, Y, Ishikawa, T & Suzuki, T 1995, 'Characterization of a chemically reactive propranolol metabolite that binds to microsomal proteins of rat liver', Chemical Research in Toxicology, vol. 8, no. 5, pp. 721-728.
Narimatsu, Shizuo ; Watanabe, Toshiyuki ; Masubuchi, Yasuhiro ; Horie, Toshiharu ; Kumagai, Yoshito ; Cho, Arthur K. ; Imaoka, Susumu ; Funae, Yoshihiko ; Ishikawa, Tsutomu ; Suzuki, Tokuji. / Characterization of a chemically reactive propranolol metabolite that binds to microsomal proteins of rat liver. In: Chemical Research in Toxicology. 1995 ; Vol. 8, No. 5. pp. 721-728.
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AU - Watanabe, Toshiyuki

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AU - Horie, Toshiharu

AU - Kumagai, Yoshito

AU - Cho, Arthur K.

AU - Imaoka, Susumu

AU - Funae, Yoshihiko

AU - Ishikawa, Tsutomu

AU - Suzuki, Tokuji

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N2 - We have characterized a chemically reactive propranolol (PL) metabolite which binds to proteins in rat liver microsomes. During incubation with rat liver microsomes (1 mg of protein) fortified with an NADPH-generating system, 4-hydroxypropranolol (4-OH-PL) quickly disappeared from the reaction medium, but none of the possible metabolite peaks was detected under the high-performance liquid chromatographic conditions used. The consumption of 4-OH-PL depended on microsomes and NADPH. The reaction was not affected by inhibitors of cytochrome P450 or FAD monooxygenase, but was markedly diminished in the presence of cytosol and ascorbic acid. The effect of cytosol was inhibited by potassium cyanide but not by sodium benzoate or dimethyl sulfoxide, and was also not affected by heating at 60°C for 30 min, suggesting that superoxide (SO) ion was involved in the reaction and that it was blocked by superoxide dismutase (SOD) present in the cytosol. Cu,Zn-SOD, purified from cytosol, effectively mimicked the suppressive effect of cytosol. Incubation of 4-OH-PL in an SO-generating system of xanthine and xanthine oxidase generated 1,4-naphthoquinone (1,4-NQ), which was identified by TLC, HPLC, and GC/MS. 1,4-NQ was also formed in microsomal incubates containing NADPH and small amounts of microsomes (below 0.1 mg of protein). These results indicate that 4-OH-PL is converted by SO, or some reactive oxygen species derived from it, to 1,4-NQ which binds to proteins and is one of the reactive metabolites of PL.

AB - We have characterized a chemically reactive propranolol (PL) metabolite which binds to proteins in rat liver microsomes. During incubation with rat liver microsomes (1 mg of protein) fortified with an NADPH-generating system, 4-hydroxypropranolol (4-OH-PL) quickly disappeared from the reaction medium, but none of the possible metabolite peaks was detected under the high-performance liquid chromatographic conditions used. The consumption of 4-OH-PL depended on microsomes and NADPH. The reaction was not affected by inhibitors of cytochrome P450 or FAD monooxygenase, but was markedly diminished in the presence of cytosol and ascorbic acid. The effect of cytosol was inhibited by potassium cyanide but not by sodium benzoate or dimethyl sulfoxide, and was also not affected by heating at 60°C for 30 min, suggesting that superoxide (SO) ion was involved in the reaction and that it was blocked by superoxide dismutase (SOD) present in the cytosol. Cu,Zn-SOD, purified from cytosol, effectively mimicked the suppressive effect of cytosol. Incubation of 4-OH-PL in an SO-generating system of xanthine and xanthine oxidase generated 1,4-naphthoquinone (1,4-NQ), which was identified by TLC, HPLC, and GC/MS. 1,4-NQ was also formed in microsomal incubates containing NADPH and small amounts of microsomes (below 0.1 mg of protein). These results indicate that 4-OH-PL is converted by SO, or some reactive oxygen species derived from it, to 1,4-NQ which binds to proteins and is one of the reactive metabolites of PL.

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