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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 fragmentation mechanism for metabolites M16 and M17.
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pone.0134027.g011: Proposed fragmentation mechanism for metabolites M16 and M17.

Mentions: M16 ([M+H]+; m/z 472.4933): The third most abundant metabolite M16 at m/z 472.4933 ([M+H]+) with an elemental composition of C6H21N4O4PtS2 (-2.96 ppm) was eluted at 12.9 min. The HRMS data suggests the lack of one chlorine atom and inclusion of an additional cysteine moiety in M16 as compared to M7. This can be seen from the LC-MS/MS spectrum of protonated M16 which shows abundant product ions at m/z 350.4819 and m/z 228.4812 corresponding to a probable loss of one and two cysteine moieties, respectively, and m/z 122.0275 (protonated cysteine). The peaks at m/z 264.4732 (Pt+(NH3)2Cl) and m/z 246.5177 (Pt+(NH3)2OH) would have been present and the sequential losses of two neutral species of cysteine would have been absent in case of one cysteine moiety in M16 as discussed in M7 and M8. The moderately abundant product ions at m/z 426.4871 (loss of HCOOH) and m/z 380.4762 (loss of two HCOOH), also designate the presence of two cysteine moieties in M16 (Fig 11). Similarly to M7 and M8, the appearance of m/z 122.0275 (protonated cysteine) and m/z 127.0576 (deuterated cysteine) in the MS/MS of protonated and deuterated M16, respectively, substantiates the presence of cysteine moiety in the structure of M16. Based on these data, M16 was characterized to be a di-cysteine metabolite of CP.


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 fragmentation mechanism for metabolites M16 and M17.
© Copyright Policy
Related In: Results  -  Collection

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

pone.0134027.g011: Proposed fragmentation mechanism for metabolites M16 and M17.
Mentions: M16 ([M+H]+; m/z 472.4933): The third most abundant metabolite M16 at m/z 472.4933 ([M+H]+) with an elemental composition of C6H21N4O4PtS2 (-2.96 ppm) was eluted at 12.9 min. The HRMS data suggests the lack of one chlorine atom and inclusion of an additional cysteine moiety in M16 as compared to M7. This can be seen from the LC-MS/MS spectrum of protonated M16 which shows abundant product ions at m/z 350.4819 and m/z 228.4812 corresponding to a probable loss of one and two cysteine moieties, respectively, and m/z 122.0275 (protonated cysteine). The peaks at m/z 264.4732 (Pt+(NH3)2Cl) and m/z 246.5177 (Pt+(NH3)2OH) would have been present and the sequential losses of two neutral species of cysteine would have been absent in case of one cysteine moiety in M16 as discussed in M7 and M8. The moderately abundant product ions at m/z 426.4871 (loss of HCOOH) and m/z 380.4762 (loss of two HCOOH), also designate the presence of two cysteine moieties in M16 (Fig 11). Similarly to M7 and M8, the appearance of m/z 122.0275 (protonated cysteine) and m/z 127.0576 (deuterated cysteine) in the MS/MS of protonated and deuterated M16, respectively, substantiates the presence of cysteine moiety in the structure of M16. Based on these data, M16 was characterized to be a di-cysteine metabolite of CP.

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