Identifying the compound formulae-related xenobiotics in bio-samples is full of challenges.Conventional strategies always exhibit the insufficiencies in overall coverage,analytical efficiency,and degree of automation,...Identifying the compound formulae-related xenobiotics in bio-samples is full of challenges.Conventional strategies always exhibit the insufficiencies in overall coverage,analytical efficiency,and degree of automation,and the results highly rely on the personal knowledge and experience.The goal of this work was to establish a software-aided approach,by integrating ultra-high performance liquid chromatography/ion-mobility quadrupole time-of-flight mass spectrometry(UHPLC/IM-QTOF-MS)and in-house high-definition MS^(2) library,to enhance the identification of prototypes and metabolites of the compound formulae in vivo,taking Sishen formula(SSF)as a template.Seven different MS2 acquisition methods were compared,which demonstrated the potency of a hybrid scan approach(namely high-definition data-independent/data-dependent acquisition(HDDIDDA))in the identification precision,MS1 coverage,and MS^(2) spectra quality.The HDDIDDA data for 55 reference compounds,four component drugs,and SSF,together with the rat bio-samples(e.g.,plasma,urine,feces,liver,and kidney),were acquired.Based on the UNIFI™platform(Waters),the efficient data processing workflows were established by combining mass defect filtering(MDF)-induced classification,diagnostic product ions(DPIs),and neutral loss filtering(NLF)-dominated structural confirmation.The high-definition MS^(2) spectral libraries,dubbed in vitro-SSF and in vivo-SSF,were elaborated,enabling the efficient and automatic identification of SSF-associated xenobiotics in diverse rat bio-samples.Consequently,118 prototypes and 206 metabolites of SSF were identified,with the identification rate reaching 80.51%and 79.61%,respectively.The metabolic pathways mainly involved the oxidation,reduction,hydrolysis,sulfation,methylation,demethylation,acetylation,glucuronidation,and the combined reactions.Conclusively,the proposed strategy can drive the identification of compound formulae-related xenobiotics in vivo in an intelligent manner.展开更多
A two-dimensional LC(2D-LC)method was coupled to an ion mobility-high-resolution mass spectrometer(IM-MS),which enables the separation of complex samples in four dimensions[2D-LC,ion mobility(IM)and mass spectrometry(...A two-dimensional LC(2D-LC)method was coupled to an ion mobility-high-resolution mass spectrometer(IM-MS),which enables the separation of complex samples in four dimensions[2D-LC,ion mobility(IM)and mass spectrometry(MS)].This approach works as a continuous multiheart-cutting LC-system,using a long modulation time of four minutes,in comparison to comprehensive two-dimensional liquid chromatography,which allows the complete transfer of most of the first dimension peaks to the second dimension column without fractionation.Hence,each compound delivers only one peak in the second dimension,which simplifies the data handling even when ion mobility as a third and mass spectrometry as a fourth dimension are introduced.The analysis of different complex samples,such as a plant extract from Hediotys diffusa and Scutellaria barbata,a waste water inflow,and a biocoal sample,was shown.The results of the four-dimensional separation method demonstrate that with the same column combination and the same solvents and gradients,that means without method optimization,totally different samples can be separated with outstanding separation power.Each sample was spiked with cyclophosphamide and ifosfamide and the ion suppression was determined by comparison of the peak area in the complex samples and in pure water after analysis of these samples with a 1D-LC and a 2D-LC approach.It is shown that the 2D-LC method allows an external calibration for the spiked compounds in the plant and waste water sample because of the higher separation power in comparison with 1D-LC.展开更多
基金This work was financially supported by National Natural Science Foundation of China(Grant No.:82192914)Tianjin Outstanding Youth Fund(Grant No.:23JCJQJC00030)the Innovation Team and Talents Cultivation Program of National Administration of Traditional Chinese Medicine(Grant No.:ZYYCXTD-C-202009).
文摘Identifying the compound formulae-related xenobiotics in bio-samples is full of challenges.Conventional strategies always exhibit the insufficiencies in overall coverage,analytical efficiency,and degree of automation,and the results highly rely on the personal knowledge and experience.The goal of this work was to establish a software-aided approach,by integrating ultra-high performance liquid chromatography/ion-mobility quadrupole time-of-flight mass spectrometry(UHPLC/IM-QTOF-MS)and in-house high-definition MS^(2) library,to enhance the identification of prototypes and metabolites of the compound formulae in vivo,taking Sishen formula(SSF)as a template.Seven different MS2 acquisition methods were compared,which demonstrated the potency of a hybrid scan approach(namely high-definition data-independent/data-dependent acquisition(HDDIDDA))in the identification precision,MS1 coverage,and MS^(2) spectra quality.The HDDIDDA data for 55 reference compounds,four component drugs,and SSF,together with the rat bio-samples(e.g.,plasma,urine,feces,liver,and kidney),were acquired.Based on the UNIFI™platform(Waters),the efficient data processing workflows were established by combining mass defect filtering(MDF)-induced classification,diagnostic product ions(DPIs),and neutral loss filtering(NLF)-dominated structural confirmation.The high-definition MS^(2) spectral libraries,dubbed in vitro-SSF and in vivo-SSF,were elaborated,enabling the efficient and automatic identification of SSF-associated xenobiotics in diverse rat bio-samples.Consequently,118 prototypes and 206 metabolites of SSF were identified,with the identification rate reaching 80.51%and 79.61%,respectively.The metabolic pathways mainly involved the oxidation,reduction,hydrolysis,sulfation,methylation,demethylation,acetylation,glucuronidation,and the combined reactions.Conclusively,the proposed strategy can drive the identification of compound formulae-related xenobiotics in vivo in an intelligent manner.
文摘A two-dimensional LC(2D-LC)method was coupled to an ion mobility-high-resolution mass spectrometer(IM-MS),which enables the separation of complex samples in four dimensions[2D-LC,ion mobility(IM)and mass spectrometry(MS)].This approach works as a continuous multiheart-cutting LC-system,using a long modulation time of four minutes,in comparison to comprehensive two-dimensional liquid chromatography,which allows the complete transfer of most of the first dimension peaks to the second dimension column without fractionation.Hence,each compound delivers only one peak in the second dimension,which simplifies the data handling even when ion mobility as a third and mass spectrometry as a fourth dimension are introduced.The analysis of different complex samples,such as a plant extract from Hediotys diffusa and Scutellaria barbata,a waste water inflow,and a biocoal sample,was shown.The results of the four-dimensional separation method demonstrate that with the same column combination and the same solvents and gradients,that means without method optimization,totally different samples can be separated with outstanding separation power.Each sample was spiked with cyclophosphamide and ifosfamide and the ion suppression was determined by comparison of the peak area in the complex samples and in pure water after analysis of these samples with a 1D-LC and a 2D-LC approach.It is shown that the 2D-LC method allows an external calibration for the spiked compounds in the plant and waste water sample because of the higher separation power in comparison with 1D-LC.
