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. 2022 Jul 18;12(7):662.
doi: 10.3390/metabo12070662.

A Comprehensive Study to Identify Major Metabolites of an Amoxicillin-Sulbactam Hybrid Molecule in Rats and Its Metabolic Pathway Using UPLC-Q-TOF-MS/MS

Fei-Ke Zhao  1 , Ren-Bin Shi  1 , Yu-Bin Sun  2 , Shuang-Yun Yang  1 , Liang-Zhu Chen  3 , Bing-Hu Fang  4
Affiliations

A Comprehensive Study to Identify Major Metabolites of an Amoxicillin-Sulbactam Hybrid Molecule in Rats and Its Metabolic Pathway Using UPLC-Q-TOF-MS/MS

Fei-Ke Zhao et al. Metabolites. .

Abstract

Amoxicillin and sulbactam are widely used compound drugs in animal food. The amoxicillin-sulbactam hybrid molecule can achieve better curative effects through the combination of the two drugs. However, its pharmacokinetic behavior needs to be explored. In this study, a randomized crossover experiment was performed to investigate the metabolism of the novel amoxicillin-sulbactam hybrid molecule in rats after gastric administration. Ultrahigh performance liquid chromatography-quadrupole time-of-flight tandem mass spectrometry (UPLC-Q-TOF-MS/MS) was used to isolate and to identify the metabolites in rats. Amoxicillin, amoxicilloic acid, amoxicillin diketopiperazine, and sulbactam were eventually detected in the plasma, liver, urine, and kidneys; no hybrid molecules and their metabolites were detected in feces. The in vivo metabolism results showed that the hybrid molecule was absorbed into the body in the intestine, producing amoxicillin and sulbactam, then amoxicillin was partially metabolized to amoxicilloic acid and amoxicillin diketopiperazine, which are eventually excreted in the urine by the kidneys. In this study, four major metabolites of the amoxicillin-sulbactam hybrid molecule were identified and their metabolic pathways were speculated, which provided scientific data for understanding the metabolism of the hybrid molecule and for its clinical rational use.

Keywords: amoxicillin; drug metabolism; hybrid molecule; sulbactam; ultrahigh performance liquid chromatography–quadrupole time-of-flight tandem mass spectrometry.

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Conflict of interest statement

The authors declare that they have no conflict of interest.

Figures

Figure A1
Figure A1
Total ion chromatograms of urine. (a,b) Blank urine; (c,d) Sample urine.
Figure A2
Figure A2
Total ion chromatograms of kidney. (a,b) Blank kidney; (c,d) Sample kidney.
Figure A3
Figure A3
Total ion chromatograms of plasma. (a,b) Blank plasma; (c,d) Sample plasma.
Figure A4
Figure A4
Total ion chromatograms of liver. (a,b) Blank liver; (c,d) Sample liver.
Figure A5
Figure A5
Total ion chromatograms of feces. (a,b) Blank feces; (c,d) Sample feces.
Figure 1
Figure 1
Chemical structure of AS.
Figure 2
Figure 2
Metabolite-prediction diagram for AS.
Figure 3
Figure 3
EIC and mass spectrometry results for AS. (a) EIC spectrometry results for AS; (b) Mass spectrum of AS; (c) Two-stage mass spectral data of AS.
Figure 4
Figure 4
EIC and mass spectrometry results for amoxicillin. (a) EIC spectrometry results for amoxicillin; (b) Mass spectrum of amoxicillin; (c) Two-stage mass spectral data of amoxicillin.
Figure 5
Figure 5
EIC and mass spectrometry results for amoxicilloic acid. (a) EIC spectrometry results for amoxicilloic acid; (b) Mass spectrum of amoxicilloic acid; (c) Two-stage mass spectral data of amoxicilloic acid.
Figure 6
Figure 6
EIC and mass spectrometry results for amoxicillin diketopiperazine. (a): EIC spectrometry results for amoxicillin diketopiperazine; (b): Mass spectrum of amoxicillin diketopiperazine; (c): Two-stage mass spectral data of amoxicillin diketopiperazine.
Figure 7
Figure 7
EIC and mass spectrometry results for Sulbactam. (a): EIC spectrometry results for sulbactam; (b): Mass spectrum of sulbactam; (c): Two-stage mass spectral data of sulbactam.
Figure 8
Figure 8
Ion chromatograms of urine samples. (a) M2; (b) M1; (c) M3; (d) M4.
Figure 9
Figure 9
Ion chromatograms of kidney samples. (a) M1; (b) M3; (c) M4.
Figure 10
Figure 10
Ion chromatograms of plasma samples. (a) M1; (b) M3; (c) M4.
Figure 11
Figure 11
Ion chromatograms of liver samples. (a) M1; (b) M3; (c) M4.
Figure 12
Figure 12
Mass spectra of four metabolites M1, M2, M3, and M4. (a) Mass spectrum of M1; (b) Two-stage mass spectral data of M1; (c) Mass spectrum of M2; (d) Two-stage mass spectral data of M2; (e) Mass spectrum of M3; (f) Two-stage mass spectral data of M3; (g) Mass spectrum of M4; (h) Two-stage mass spectral data of M4.
Figure 13
Figure 13
Possible structures of fragment ions in amoxicillin. (a) Possible structure of the m/z 349 fragment ion; (b) Possible structure of the m/z 208 fragment ion; (c) Possible structure of the m/z 160 fragment ion; (d) Possible structure of the m/z 114 fragment ion.
Figure 14
Figure 14
Two possible positions of fragment m/z 323 ions in amoxicilloic acid.
Figure 15
Figure 15
Two possible positions of fragment m/z 189 ions in amoxicilloic acid.
Figure 16
Figure 16
Possible structure of fragment ions in amoxicillin diketopiperazine. (a) Possible structure of fragment m/z 207 ion; (b) Possible structure of fragment m/z 160 ion.
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