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
Herbal components characterization represents a challenging task because of the co-existing of multiple classes of naturally occurring compounds with wide spans of polarity,molecular mass,and the ubiquitous isomerism....Herbal components characterization represents a challenging task because of the co-existing of multiple classes of naturally occurring compounds with wide spans of polarity,molecular mass,and the ubiquitous isomerism.The root and rhizome of Salvia miltiorrhiza have been utilized as a reputable traditional Chinese medicine Salviae Miltiorrhizae Radix et Rhizoma(Dan-Shen)in the treatment of cardiovascular disease.Herein,a dimensionenhanced ultra-high performance liquid chromatography/ion mobility/quadrupole time-of-flight mass spectrometry approach in combination with intelligent peak annotation workflows was established aimed to rapidly characterize the multicomponents from S.miltiorrhiza.Due to the sufficient optimization,satisfactory chromatography separation was enabled on an HSS T3 column within 33 min using 0.1%formic acid in water(A)and acetonitrile(B)as the mobile phase,while the data-independent HDMS^(E) in both the negative and positive electrospray ionization modes was utilized for the high-coverage MS^(2) data acquisition.Streamlined automatic peak annotation by searching an in-house library(recording 198 known compounds)followed by the subsequent confirming steps(e.g.,comparison with the reference compounds,fragmentation pathways analysis,and retention behavior comparison,etc.),allowed us to identify or tentatively characterize a total of 86 components(including 50 terpenoids,21 phenolic acids,and 15 others)from S.miltiorrhiza.Importantly,three-dimensional structure information,such as the retention time,MS^(1) and MS^(2) data,and collision cross section(CCS),was provided,which can facilitate the more reliable characterization of herbal components.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.
基金This work was financially supported by the National Key Research and Development Program of China(Grant No.2018YFC1704500)Tianjin Committee of Science and Technology of China(Grant No.21ZYJDJC00080)National Natural Science Foundation of China(Grant No.81872996)。
文摘Herbal components characterization represents a challenging task because of the co-existing of multiple classes of naturally occurring compounds with wide spans of polarity,molecular mass,and the ubiquitous isomerism.The root and rhizome of Salvia miltiorrhiza have been utilized as a reputable traditional Chinese medicine Salviae Miltiorrhizae Radix et Rhizoma(Dan-Shen)in the treatment of cardiovascular disease.Herein,a dimensionenhanced ultra-high performance liquid chromatography/ion mobility/quadrupole time-of-flight mass spectrometry approach in combination with intelligent peak annotation workflows was established aimed to rapidly characterize the multicomponents from S.miltiorrhiza.Due to the sufficient optimization,satisfactory chromatography separation was enabled on an HSS T3 column within 33 min using 0.1%formic acid in water(A)and acetonitrile(B)as the mobile phase,while the data-independent HDMS^(E) in both the negative and positive electrospray ionization modes was utilized for the high-coverage MS^(2) data acquisition.Streamlined automatic peak annotation by searching an in-house library(recording 198 known compounds)followed by the subsequent confirming steps(e.g.,comparison with the reference compounds,fragmentation pathways analysis,and retention behavior comparison,etc.),allowed us to identify or tentatively characterize a total of 86 components(including 50 terpenoids,21 phenolic acids,and 15 others)from S.miltiorrhiza.Importantly,three-dimensional structure information,such as the retention time,MS^(1) and MS^(2) data,and collision cross section(CCS),was provided,which can facilitate the more reliable characterization of herbal components.
