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Liquid Chromatography Electrospray Ionization Tandem Mass Spectrometric (LC/ESI-MS/MS) Study for the Identification and Characterization of In Vivo Metabolites of Cisplatin in Rat Kidney Cancer Tissues: Online Hydrogen/Deuterium (H/D) Exchange Study.

Bandu R, Ahn HS, Lee JW, Kim YW, Choi SH, Kim HJ, Kim KP - PLoS ONE (2015)

Bottom Line: A total of thirty one unknown in vivo metabolites have been identified and the structures of metabolites were elucidated using LC-MS/MS experiments combined with accurate mass measurements.Online HDX experiments have been used to further support the structural characterization of metabolites.The results showed that CP undergoes a series of ligand exchange biotransformation reactions with water and other nucleophiles like thio groups of methionine, cysteine, acetylcysteine, glutathione and thioether.

View Article: PubMed Central - PubMed

Affiliation: Department of Applied Chemistry, College of Applied Sciences, Kyung Hee University, Yong-in City, Republic of Korea.

ABSTRACT
In vivo rat kidney tissue metabolites of an anticancer drug, cisplatin (cis-diamminedichloroplatinum [II]) (CP) which is used for the treatment of testicular, ovarian, bladder, cervical, esophageal, small cell lung, head and neck cancers, have been identified and characterized by using liquid chromatography positive ion electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS) in combination with on line hydrogen/deuterium exchange (HDX) experiments. To identify in vivo metabolites, kidney tissues were collected after intravenous administration of CP to adult male Sprague-Dawley rats (n = 3 per group). The tissue samples were homogenized and extracted using newly optimized metabolite extraction procedure which involves liquid extraction with phosphate buffer containing ethyl acetate and protein precipitation with mixed solvents of methanol-water-chloroform followed by solid-phase clean-up procedure on Oasis HLB 3cc cartridges and then subjected to LC/ESI-HRMS analysis. A total of thirty one unknown in vivo metabolites have been identified and the structures of metabolites were elucidated using LC-MS/MS experiments combined with accurate mass measurements. Online HDX experiments have been used to further support the structural characterization of metabolites. The results showed that CP undergoes a series of ligand exchange biotransformation reactions with water and other nucleophiles like thio groups of methionine, cysteine, acetylcysteine, glutathione and thioether. This is the first research approach focused on the structure elucidation of biotransformation products of CP in rats, and the identification of metabolites provides essential information for further pharmacological and clinical studies of CP, and may also be useful to develop various effective new anticancer agents.

No MeSH data available.


Related in: MedlinePlus

Proposed in vivo ligand exchange biotransformation pathway of CP.
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pone.0134027.g002: Proposed in vivo ligand exchange biotransformation pathway of CP.

Mentions: Fig 1 shows the LC/ESI-MS-TIC of metabolites of CP formed in kidney tissue homogenate samples. All the metabolites (M1-M31) were eluted within 35 min on C-18 column and they were well resolved from parent drug as well as from each other. The positive ion ESI-MS spectrum of CP gives moderately abundant [M+H]+, [M+NH4]+ ions and abundant [M+Na]+ ion, and low abundance [2M+H]+, [2M+NH4]+ [2M+Na]+ ions, whereas the positive ion ESI-MS spectra of all metabolites give abundant [M+H]+ ions and moderately abundant [M+Na]+ ions. To elucidate the structures of metabolites, we examined the fragmentation of the [M+H]+ ions of CP and its metabolites by using online LC/ESI-MS/MS experiments in combination with accurate mass measurements. Further, online HDX LC-MS/MS experiments were also carried out to support the structural identification of metabolites. The product ions of all the metabolites were compared with the product ions of CP, to assign most probable structures for the observed metabolites (Fig 2). The elemental compositions of thirty one metabolites and their majority of product ions have been confirmed by accurate mass measurements data.


Liquid Chromatography Electrospray Ionization Tandem Mass Spectrometric (LC/ESI-MS/MS) Study for the Identification and Characterization of In Vivo Metabolites of Cisplatin in Rat Kidney Cancer Tissues: Online Hydrogen/Deuterium (H/D) Exchange Study.

Bandu R, Ahn HS, Lee JW, Kim YW, Choi SH, Kim HJ, Kim KP - PLoS ONE (2015)

Proposed in vivo ligand exchange biotransformation pathway of CP.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4526507&req=5

pone.0134027.g002: Proposed in vivo ligand exchange biotransformation pathway of CP.
Mentions: Fig 1 shows the LC/ESI-MS-TIC of metabolites of CP formed in kidney tissue homogenate samples. All the metabolites (M1-M31) were eluted within 35 min on C-18 column and they were well resolved from parent drug as well as from each other. The positive ion ESI-MS spectrum of CP gives moderately abundant [M+H]+, [M+NH4]+ ions and abundant [M+Na]+ ion, and low abundance [2M+H]+, [2M+NH4]+ [2M+Na]+ ions, whereas the positive ion ESI-MS spectra of all metabolites give abundant [M+H]+ ions and moderately abundant [M+Na]+ ions. To elucidate the structures of metabolites, we examined the fragmentation of the [M+H]+ ions of CP and its metabolites by using online LC/ESI-MS/MS experiments in combination with accurate mass measurements. Further, online HDX LC-MS/MS experiments were also carried out to support the structural identification of metabolites. The product ions of all the metabolites were compared with the product ions of CP, to assign most probable structures for the observed metabolites (Fig 2). The elemental compositions of thirty one metabolites and their majority of product ions have been confirmed by accurate mass measurements data.

Bottom Line: A total of thirty one unknown in vivo metabolites have been identified and the structures of metabolites were elucidated using LC-MS/MS experiments combined with accurate mass measurements.Online HDX experiments have been used to further support the structural characterization of metabolites.The results showed that CP undergoes a series of ligand exchange biotransformation reactions with water and other nucleophiles like thio groups of methionine, cysteine, acetylcysteine, glutathione and thioether.

View Article: PubMed Central - PubMed

Affiliation: Department of Applied Chemistry, College of Applied Sciences, Kyung Hee University, Yong-in City, Republic of Korea.

ABSTRACT
In vivo rat kidney tissue metabolites of an anticancer drug, cisplatin (cis-diamminedichloroplatinum [II]) (CP) which is used for the treatment of testicular, ovarian, bladder, cervical, esophageal, small cell lung, head and neck cancers, have been identified and characterized by using liquid chromatography positive ion electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS) in combination with on line hydrogen/deuterium exchange (HDX) experiments. To identify in vivo metabolites, kidney tissues were collected after intravenous administration of CP to adult male Sprague-Dawley rats (n = 3 per group). The tissue samples were homogenized and extracted using newly optimized metabolite extraction procedure which involves liquid extraction with phosphate buffer containing ethyl acetate and protein precipitation with mixed solvents of methanol-water-chloroform followed by solid-phase clean-up procedure on Oasis HLB 3cc cartridges and then subjected to LC/ESI-HRMS analysis. A total of thirty one unknown in vivo metabolites have been identified and the structures of metabolites were elucidated using LC-MS/MS experiments combined with accurate mass measurements. Online HDX experiments have been used to further support the structural characterization of metabolites. The results showed that CP undergoes a series of ligand exchange biotransformation reactions with water and other nucleophiles like thio groups of methionine, cysteine, acetylcysteine, glutathione and thioether. This is the first research approach focused on the structure elucidation of biotransformation products of CP in rats, and the identification of metabolites provides essential information for further pharmacological and clinical studies of CP, and may also be useful to develop various effective new anticancer agents.

No MeSH data available.


Related in: MedlinePlus