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Matrine (MT) is a major bioactive compound extracted from

Along with definite clinical effects, traditional Chinese medicine (TCM) has increasingly gained worldwide attention. Quality control marker (Q-marker) is always used to confirm the primary effects of herbs or TCM. Thus, the characterization of the metabolism features and subsequently induced effects are essential to reveal the functional consequences. In this work, a strategy by integrating metabolite profiling of a Q-marker and network pharmacology was proposed and applied to explore the potential function of herbs, and prim-O-glucosylcimifugin, the Q-marker of Radix Saposhnikoviae, was used as an example. As a result, 35 metabolites were characterized in rats' plasma, urine, feces, heart, liver, spleen, lung, kidney, and brain. Among them, M13/M17/M20/M23/M24/M28 were the primary metabolites, and 23 metabolites were reported for the first time. It was found that major metabolic pathways of prim-O-glucosylcimifugin were hydroxylation, dehydrogenation, hydrogenation and glucuronidation. In addition to the above metabolic reactions, methylation, hydrogenation, glycosylation, isomerization, sulfation, and other S-conjunctions were found in prim-O-glucosylcimifugin for the first time. Meanwhile, targets of prim-O-glucosylcimifugin and its major metabolites targeted additional 125 targets with functions of MAPK signaling pathway, proteoglycans in cancer, pathways in cancer, bladder cancer, colorectal cancer, serotonergic synapse and natural killer cell-mediated cytotoxicity. Among them, the depression of inflammation by MAPK signaling pathway might be the core mechanism for prim-O-glucosylcimifugin to treat respiratory tract infections. The above results provided vital information for understanding the metabolism and functional mechanism of prim-O-glucosylcimifugin in the treatment of respiratory tract infections, and a new insight for revealing the pharmacological mechanism of the complex system was also provided.

Food additives are widely used in our daily life, and the side-effects caused by them have gained extensive attention around the world. Notably, constituent-oriented metabolites, in some sense, always contribute to pharmacological changes, inducing toxicity, therapeutic effects, etc. Characterization of the metabolites and their potential functions is of great importance to the practical applications. In this work, an integrated strategy by combining metabolite profiling and network pharmacology was applied to characterize the metabolic features and reveal pharmacological changes of neohesperidin dihydrochalcone (NHDC) in vivo to demonstrate its pharmacological mechanism and potential functions. As a result, a total of 19 metabolites (3 in plasma, 19 in urine, 8 in feces, 3 in heart, 5 in liver, 0 in spleen, 1 in lung, 2 in kidneys and 2 in brain) were screened and 18 of them were characterized for the first time. Phase I metabolic reactions of hydrolysis and phase II reactions of glucuronidation, sulfation, glutamylation, N-butyryl glycylation and lactylation were the main metabolic reactions of NHDC in vivo. Moreover, the results analyzed by network pharmacology revealed that, in addition to common pathways (steroid hormone biosynthesis) of NHDC, metabolites' targets were involved in pathways in cancer, ovarian steroidogenesis, proteoglycans in cancer, PI3K-Akt signaling pathway and progesterone-mediated oocyte maturation, indicating that these functional changes might result in potential novel functions or other side-effects, such as a disorder of steroid hormones. Our work provided the metabolic features and functional modifications of NHDC in vivo for the first time, and meaningful information for further pharmacological validations or potential functions is supplied.

Elucidating the therapeutical basis and functional mechanism of traditional Chinese medicine (TCM) is still a challenge faced by researchers since the effects of TCM are always achieved by the interactions of multiple components and multiple targets. In this study, an integrated strategy using a combination of chemical profiling, network pharmacology and in vitro experimental validation was applied to profile the trace constituents of TCM and characterize their functional mechanism, and Rhizoma coptidis (RC) was used as an example. As a result, a total of 55 compounds were identified or tentatively characterized in the water extract of RC, including 22 organic acids, 17 alkaloids, 8 lignans, 2 flavonoids, 4 phenylethanoid glycosides, and 2 other types of compounds. Meanwhile, a potential novel compound (compound 41) – one of the alkaloids – was found and characterized. Notably, organic acids in RC were systematically characterized for the first time. Moreover, the pharmacological targets of the 46 main constituents of RC were predicted using the network pharmacological method, and 234 of them were involved in diabetes mellitus (DM). KEGG analysis indicated that the Rap1 signaling pathway, the cAMP signaling pathway, the PI3K–Akt signaling pathway, and inflammatory mediator regulation of TRP channels were the core signaling pathways for RC in the treatment of DM. Further, the anti-inflammatory effects of RC, correlated with inflammatory mediator regulation of TRP channels, were confirmed by using the LPS-induced RAW 264.7 cell model. Meanwhile, the downstream pathway (NF-kB) of TRP channels was validated by western blot. Our work comprehensively provided the material basis and potential pharmacological mechanism of RC against inflammation in DM; in the meantime, a new insight for elucidating the functional mechanism of TCM was also supplied.

Two new xanthones, oxisterigmatocystins J and K (1–2), and two new anthraquinones, versicolorins D and E (3–4), were isolated from solid cultures of the fungus Penicillium sp. DWS10-P-6, together with twelve known compounds (5–16). Their structures, including their absolute configurations, were characterized on the basis of extensive 1D NMR, 2D NMR, MS and CD spectral data. The cytotoxic activities of compounds 1–12 against HL-60, MDA-MB-231 and PC-3 cells were also evaluated. Compounds 4 and 5 showed significant cytotoxic activity against the HL-60 cell line with IC50 values of 1.65 μM and 1.05 μM, respectively.

Objectives This study was designed to profile the chemical information of Forsythiae Fructus (FF) and investigate the in-vivo FF-related xenobiotics, especially for lignans. Methods Rats were oral administrated of FF and pinoresinol-4-O-glucoside, respectively. Blood and urine samples were collected after ingestion, and xenobiotics was profiled by an UPLC/Qtof MS method. Key findings A total of 19 lignans were identified or tentatively characterized in FF, and 63 lignan-related xenobiotics were found in rat plasma and urine after ingestion of FF. It was found that lignans could be transformed into metabolites by furan ring opening, hydrogenation, demethylation, dehydration and phase II reactions (sulfation and glucuronidation). The whole metabolic behaviour of bisepoxylignan was revealed by evaluating the metabolism of pinoresinol-4-O-glucoside in vivo. It was found that the configuration of C-8/C-8ʹ was retained after furan ring opening and metabolic reactions always occurred at position of C-3/C-4/C-5 or C-3ʹ/C-4ʹ/C-5ʹ. Additionally, other types components in FF and in vivo were also characterized. Conclusions This work revealed the in-vivo metabolism of FF, and reported the characteristic metabolic reactions of lignans for the first time. It was also provided the foundation for the further investigation on pharmacodynamic components of FF or TCMs containing FF.

RATIONALE Salvianolic acid B (Sal B), the Q-marker in Salvia miltiorrhiza, was proved to present an obvious anti-diabetes effect when treating as a food intake. Till now, its metabolism feature, tissue distribution, and anti-diabetes mechanism of Sal B were not fully elucidated. METHODS The metabolites of Sal B in rats were profiled by an ultra-high-performance liquid chromatography coupled time-of-flight mass spectrometry (UHPLC/Q-TOF MS) method. The potential anti-diabetes mechanism of Sal B was predicted by network pharmacology. RESULTS A total of 31 metabolites were characterized in rats after ingestion of Sal B at a dosage of 40 mg/kg, including 1 in plasma, 19 in urine, 31 in feces, 0 in heart, 0 in liver, 0 in spleen, 1 in lung, 1 in kidney and 0 in brain. Among them, 18 metabolites were reported for the first time. Phase I reactions of hydrolysis, hydrogenation, dehydroxylation, hydroxylation, decarboxylation, isomerization, and phase II reactions of methylation were found in Sal B. Notably, decarboxylation and dehydroxylation were revealed in salvianolic acid B for the first time. The pharmacology network results showed that Sal B and its metabolites could regulate ALB, PLG, ACE, CASP3, MMP9, MMP2, and MTOR, etc. The above targets were involved in insulin signaling pathway, PI3K-Akt signaling pathway, HIF-1 signaling pathway, and TNF signaling pathway, etc. CONCLUSION: The metabolism feature of Sal B in vivo was systematically revealed, and its anti-diabetes mechanism for further pharmacological validations was predicted based on metabolite profiling and network pharmacology for the first time.

Background Quality control exerted great importance on the clinical application of drugs for ensuring effectiveness and safety. Due to chemical complexity, diversity among different producing areas and harvest seasons, as well as unintentionally mixed with non-medicinal parts, the current quality standards of traditional Chinese medicine (TCM) still faced challenges in evaluating the overall chemical consistency. Purpose We aimed to develop a new strategy to discover potential quality marker (Q-marker) of TCM by integrating plant metabolomics and network pharmacology, using Periplocae Cortex (GP, the dried root bark of Periploca sepium Bge.) as an example. Methods First, plant metabolomics analysis was performed by UPLC/Q-TOF MS in 89 batches of samples to discover chemical markers to distinguish medicinal parts (GP) and non-medicinal parts (the dried stem bark of Periploca sepium Bge. (JP)), harvest seasons and producing region of Periplocae Cortex. Second, network pharmacology was applied to explore the initial linkages among chemical constituents, targets and diseases. Last, potential Q-marker were selected by integrating analysis of plant metabolomics and network pharmacology, and the quantification method of Q-marker was developed by using UPLC-TQ-MS. Results The chemical profiling of GP and JP was investigated. Fifteen distinguishing features were designated as core chemical markers to distinguish GP and JP. Besides, the content of 4-methoxybenzaldehyde-2-O-β- d -xylopyranosyl-(1→6)-β- d -glucopyranoside could be used to identify Periplocae Cortex harvested in spring-autumn or summer. Meanwhile, a total of 15 components targeted rheumatoid arthritis were screened out based on network pharmacology. Taking absorbed constituents into consideration, 23 constituents were selected as potential Q-marker. A simultaneous quantification method (together with 11 semi-quantitative analysis) was developed and applied to the analysis of 20 batches of commercial Periplocae Cortex on the market. The PLS-DA model was successfully developed to distinguish GP and JP samples. In addition, the artificially mixed GP sample, which contained no less than 10% of the adulterant (JP), could also be correctly identified. Conclusion Our results indicated that 9 ingredients could be considered as Q-marker of Periplocae Cortex. This study has also demonstrated that the plant metabolomics and network pharmacology could be used as an effective approach for discovering Q-marker of TCM to fulfill the evaluation of overall chemical consistency among samples from different producing areas, harvest seasons, and even those commercial crude drugs, which might be mixed with a small amount of non-medicinal parts.

Ethnopharmacological relevance Shuang-Huang-Lian preparation has captured wide attention since its clinical applications for the successful treatment of upper respiratory tract infection. However, its functional basis under actual therapeutic dose in vivo was still unrevealed. Aim of the study This study aimed to reveal the anti-flu substances and mechanism of Shuang-Huang-Lian water extract (SHL) on H1N1 infected mouse model by a strategy based on serum pharmaco-chemistry under actual therapeutic dose and network pharmacology. Materials and methods H1N1 infected mouse model was employed for evaluation of the anti-flu effects of SHL. A simultaneous quantification method was developed by UPLC-TQ-XS MS coupled switch-ions mode and applied to characterize the pharmacokinetics of the multiple components of SHL under actual therapeutic dose. The potential active ingredients were screened out based on their pharmacokinetic parameters. And then, a compound mixture of these active candidates was re-evaluated for the anti-flu activity on H1N1 infected mouse model. Furthermore, the anti-flu mechanism of SHL was also predicted by network pharmacology coupled with the experimental result. Results SHL significantly increased the survival rate and prolonged survival days on H1N1 infected mice at a dosage of 20 g crude drug/kg/day by reversing the increased lung index, down-regulating the inflammatory cytokines (TNF-α, IL-1β, IL-6) and inhibiting the release of IFN-β in bronchoalveolar lavage fluids (BALF). Concomitantly, the pharmacokinetic parameters of fourteen quantified and twenty-one semi-quantified constituents of SHL were characterized. And then, five compounds (baicalin, sweroside, chlorogenic acid, forsythoside A and phillyrin), which displayed satisfactory pharmacokinetic features, were considered as potential active ingredients. Thus, a mixture of these five ingredients was administered to H1N1-infected mice at a dose of 4.24 mg/kg/day. As a result, the therapeutical effects of the mixture were similar to SHL in terms of survival rate, lung index and the release of cytokines (TNF-α, IL-1β and IL-6) in BALF. Moreover, network pharmacology analysis indicated that the TNF-signal pathways might play a role in the anti-flu mechanism of SHL. Conclusions A mixture of five compounds (baicalin, sweroside, chlorogenic acid, forsythoside A and phillyrin) were the anti-flu substances of SHL. The strategy based on serum pharmaco-chemistry under actual therapeutic dose provided a new sight on exploring in vivo effective substances of TCM.

Two new xanthones, oxisterigmatocystins J and K (1-2), and two new anthraquinones, versicolorins D and E (3-4), were isolated from solid cultures of the fungus

Rationale Traditional Chinese medicines (TCMs) attract worldwide attention because of their effects in clinical application recorded in China historical ancient codes and in records, such as 'Treatise on Febrile Diseases'. With the developments of drug analysis and research, evaluating the in vivo substances in TCMs has become of great importance. Scutellariae Radix (SR, named as huang-qing in China), the root of Scutellaria baicalensis Georgi, has shown favorable clinical effects and safety in the treatment of infection diseases; however, its in vivo compounds are unclear and need detailed investigation. Methods An ultra-high-performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UHPLC/QTOF MS) method coupled to an integrated strategy involving diagnostic ions, neutral losses and a prediction platform was used to explore the constituents of SR, and their exogenous substances in rats. Results A total of 118 chemical constituents mainly featuring five chemical structure types (flavone C-glycosides, flavone O-glycosides, free flavones, flavanones and phenylethanoid glycosides) were identified or tentatively characterized in SR, and 175 xenobiotics (68 prototypes and 107 metabolites) were profiled in rat plasma, urine, bile and feces after ingestion of SR. The metabolites were classified into four related chemical groups: flavone C-glycosides, flavone O-glycosides, flavanones and phenylethanoid glycosides. Phase II metabolism reactions, such as glucuronidation and sulfation, were the major metabolic reactions in addition to phase I reactions of hydrolysis and hydrogenation. The corresponding main metabolic features of SR in rats were also elucidated. Conclusions The metabolism of SR, as a whole, was systemically revealed for the first time, and our work also provided meaningful information for pharmacokinetics studies and pharmacological analysis of SR in future work.

Lonicerae japonicae flos.(LJF) was widely used as a drug to treat upper respiratory tract infection or a tea to clear heat in Asian countries for thousands of years. Despite of its curative effects confirmed by modern pharmacological methods, its functional materials and mechanism against influenza were still unclear and needed further investigation. In this study, an integrated strategy based on in vivo substances profiling and network pharmacology was proposed and applied to screen out the potential anti-influenza substances and mechanism of LJF. An UHPLC/Q-TOF MS method was utilized to profile the chemical components in LJF and their metabolites in rats. The targets of absorbed prototypes were predicted by Swiss Target Prediction, and they were further analyzed by String and Kyoto Encyclopedia of Genes and Genomes (KEGG). As a result, a total of 126 chemical components mainly featuring three chemical structure types were characterized, including 70 iridoid glycosides, 17 caffeoylquinic acids, 24 flavonoids, and 15 other types compounds. Among them, ten N-contained iridoid glycosides were characterized as potential novel compounds. Moreover, 141 xenobiotics (74 prototypes and 67 metabolites) were clearly screened out in rat plasma and urine after ingestion of LJF. Phase II reactions (sulfation, glucuronidation, methylation) and phase I reactions (dehydroxylation, hydrogenation, hydrolysis, N-heterocyclization) were the main metabolic reactions of LJF in rats. Further, a total of 338 targets were predicted and TNF, PTGS2 and EGFR were the three main targets involved in the pathology of influenza. In addition to normal NF-κB pathway, T cell signal pathway and mTOR signal pathway were the other patterns for LJF to achieve its anti-flu effects. Our work provided the meaningful data for further pharmacological validation of LJF against influenza, and a new strategy was also proposed for minimizing the process to reveal the mechanism and functional basis of TCMs.

Traditional Chinese medicine was widely used in China since its definite effects and therapy. The components of TCM were absorbed into the circle system as the format of prototypes or metabolites, which contributed to the therapy or side effects. Declaring the functional changes in this process was of great importance to the clinical applications. In this work, an integrated strategy based on metabolites’ profiling and network pharmacology was proposed for exploring the pharmacological changes of compounds in vivo. Arctiin, the main component in Fructus Arctii with various kinds of bioactivities, was used as an example. An ultra-performance liquid chromatography coupled with time-of-flight mass spectrometry and metabolynx™ software was applied to characterize the metabolites of arctiin in rats at a dosage of 100 mg/kg; network pharmacology was applied to characterize the functional changes. As a result, fifty-three metabolites (32 in plasma, 40 in urine, 19 in bile, 20 in feces, 1 in brain, 12 in liver and 4 in lungs) were screened out and characterized, and 3 of them were unambitiously identified by comparison with standard substances. Among them, 38 metabolites were reported for the first time. It was found the major metabolic pathways of arctiin in rats were demethylation, lactone-opening and phase II conjugations with sulfate and glucuronide. It also confirmed that M14, M15, M18, M23, M22, M43 and M45 were the major circulating forms of arctiin in rats following oral administration. In addition to the above metabolic reactions, phase I reactions of hydrolysis, demethylation, dehydroxylation were also observed, and dehydrogenation were first revealed metabolic patterns of arctiin in rats. Meanwhile, in addition to the main targets of arctiin (MTOR, EGFR and MAPK14), its metabolites targeted additional 392 targets with additional functions of anti-hepatitis B or viral carcinogenesis (SRC, CAPS3, PIK3CA, CDK4, ESR1, MMP9 and ERBB2). The above results provided very important information for understanding the metabolism and functional changes of arctiin in vivo, and supporting data for further pharmacological evaluation. Our work also provided a newsight for elucidation of functional changes of TCMs in vivo.

Traditional Chinese medicine (TCM) is regarded as a large database containing hundreds to thousands of chemical constituents that can be further developed as clinical drugs, such as artemisinin in Artemisia annua. However, effectively exploring novel candidates is still a challenge faced by researchers.In this work, an integrated strategy combining chemical profiling, molecular networking, chemical isolation, and activity evaluation (CMCA strategy) was proposed and applied to systematically characterize and screen novel candidates, and Forsythiae fructus (FF) was used as an example.It contained four parts. First, the chemical compounds in FF were detected by ultra-high-performance liquid chromatography-mass spectrometry (UPLC/Q-TOF MS) with data-dependent acquisition, and further, the targeted compounds were screened out based on an in-house database. In the meantime, the representative MS/MS fragmentation behaviors of different chemical structure types were summarized. Second, homologous constituents were grouped and organized based on feature-guided molecular networking, and the nontargeted components with homologous mass fragmentation behaviors were characterized. Third, the novel compounds were isolated and unambiguously identified by nuclear magnetic resonance (NMR). Finally, the anti-angiotensin-converting enzyme 2 (ACE2) activities of isolated chemical constituents were further evaluated by in vitro experiments.A total of 278 compounds were profiled in FF, including 151 targeted compounds and 127 nontargeted compounds. Among them, 16 were unambitiously identified by comparison with reference standards. Moreover, 25 were classified into potential novel compounds. Two novel compounds were unambiguously identified by using conventional chromatographic methods, and they were named phillyrigeninside D (peak 254) and forsythenside O (peak 155). Furthermore, the ACE2 activity of the compounds in FF was evaluated by modern pharmacological methods, and among them, suspensaside A was confirmed to present obvious anti-ACE2 activity.Our work provides meaningful information for revealing potential FF candidates for the treatment of COVID-19, along with new insight for exploring novel candidates from complex systems.

Periplocae Cortex, named Xiang-Jia-Pi in China, has been widely used to treat autoimmune diseases, especially rheumatoid arthritis. However, the in vivo substances of Periplocae Cortex remain unknown yet. In this study, an ultra-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry was used for profiling the chemical components and related metabolites of Periplocae Cortex. A total of 98 constituents were identified or tentatively characterized in Periplocae Cortex: 42 C21 steroidal glycosides, 10 cardiac glycosides, 23 organic acids, 4 aldehydes, 7 triterpenes, and 12 other types. Among them, 18 components were unambiguously identified by comparison with reference standards. In addition, 176 related xenobiotics (34 prototypes and 142 metabolites) were screened out and characterized in rats' biosamples (plasma, urine, bile, and feces) after the oral administration of Periplocae Cortex. Moreover, the metabolic fate of periplocoside S-4a, a C21 steroidal glycoside, was proposed for the first time. In summary, phase II reactions (methylation, glucuronidation, and sulfation), phase I reactions (hydrolysis reactions, oxygenation, and reduction), and their combinations were the predominant metabolic reactions of Periplocae Cortex in rat. It is the first report to reveal the in vivo substances and metabolism feature of Periplocae Cortex. This study also provided meaningful information for further pharmacodynamics study of Periplocae Cortex, as well as its quality control research.

Ziziphi Spinosae Semen (ZSS), the seeds of Ziziphus jujuba var. spinosa, is widely used in China or other Asian countries for the treatment of insomnia and palpitation. In our previous work, chemical constituents in ZSS were profiled by ultra-high-performance liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry (UHPLC/Q-TOF MS). Notably, characterization of substances in vivo was of great importance to reveal the therapy basis or mechanism in further work. Till now, there were few reports about in vivo substances' investigation of ZSS. In the present study, an integrated strategy contained represented compounds and diagnostic ions extraction was applied to characterize metabolism feature of ZSS in rats based on UHPLC/Q-TOF MS method. First, the metabolic information of four compounds (spinosin, isovitexin, jujuboside B, betulinic acid) featuring three representative chemical structures (flavonoids, saponins, terpenes) in ZSS was conducted, and their metabolism features were summarized, especially for flavonoid C-glycosides. Second, the absorbed compounds and representative compounds-related metabolites were quickly screened out; during this time, the diagnostic ions were sorted out. Last, with the help of diagnostic ions and summarized metabolic reactions, other metabolites were characterized. As a result, a total of 151 xenobiotics (58 prototypes and 93 metabolites) were identified or tentatively characterized in rats after ingestion of ZSS. Among them, 16 substances were presented in plasma, 114 in urine, 51 in bile, and 120 in feces, respectively. Hydrogenation, hydrolysis, and glucuronidation were the major metabolic reactions of ZSS in rats. The present study provided meaningful data for further pharmacological mechanism research or pharmacokinetics evaluation of ZSS.

A strategy for rapid identification of target and non-target components from traditional Chinese medicines (TCMs) extracts were proposed by utilizing the UNIFI informatics platform for the computer-assisted UPLC/Qtof MS data analyses. Ziziphi Spinosae Semen (ZSS) contains various bioactive chemical ingredients, such as flavonoids, saponins, alkaloids and terpenes. Currently, there is no method that allows rapid and comprehensive identification of these multiple components. The rapid identification of chemical components in ZSS was successfully achieved with this strategy. As a result, 60 target components were identified and 53 non-target components were characterized. Among them, chemical structures of 40 new components were deduced based on their characteristic MS fragmentation patterns. In addition, the chemical ingredients of Ziziphi Mauritianae Semen (ZMS), which is often used as substitution of ZSS, were also investigated with the same strategy. A total of 132 chemical components were identified from these two plants, including 7 additional non-target new components. It demonstrated that this strategy not only facilitated an efficient protocol for the screening and identification of target components, but also offered a new perspective on discovering non-target components in TCMs or other herbal medicines. Furthermore, 48 components were selected for semi-quantitative analyses to evaluate the difference in chemical ingredients between these two seeds of Ziziphus species. The results showed that ZSS enriched many saponins, while ZMS contained few saponins. On the contrary, many cyclopeptide alkaloids could be detected in ZMS with high content, but rare in ZSS. These results can be used for the differentiation between ZSS and its adulterant (ZMS), and also to set a scientific foundation for the establishment of quality control of ZSS.

This papercarries out a survey of sufficient schedulability analysis forfixed priority (FP) scheduling. The most common used fixed priority assignment is the rate monotonic (RM) algorithm, according to its policy, the task priorities are ordered based on their activation rates, so that the task with the shortest period is assigned the highest priority. However, when each task’s relative deadline is not equal to its period, the RM algorithm is not suitable to assign the task priorities. When relative deadlines are less than or equal to periods,the deadline monotonic (DM) algorithm can be deployed to schedule the tasks. The utilization based schedulability analysis has the advantage of simple implementation, and manyexisting schedulability analyses on uniprocessor are covered.

Traditional Chinese medicines (TCMs) usually contained a large number of chemical components, which could be transformed into more complex metabolites in vivo. In this work, a "Representative compounds-Metabolism platform-Diagnostic extraction" strategy (RMD strategy) was proposed for comprehensively identification or characterization of xenobiotics in rat after oral administration of TCMs. Shuang-Huang-Lian oral liquid (SHL), a well-known traditional Chinese medicine preparation, was used as an example. The metabolic pathways of six representative compounds, bearing five different core structures in SHL, were elucidated and their metabolic reactions were employed for exploring metabolites of homologous components in metabolism platform. Meanwhile, diagnostic ions extraction were also used for screening more structural analogues in biofluids. All this work was completed by ultra-performance liquid chromatography coupled electrospray ionization quadruple time-of-flight mass spectrometry (UPLC/Qtof MS) and UNIFI metabolism platform. As a result, a total of 254 xenobiotics were identified or tentatively characterized in rat plasma and urine after oral administration of SHL and six representative compounds. The metabolism reaction included phase I reaction (hydroxylation, hydrolysis reaction, deglycosylation, hydrogenation, demethylation, dehydroxylation and ring opening reaction) and phase II reaction (glucuronidation, sulfation and methylation). This research provided useful information for further study of the pharmacology and mechanism of SHL in vivo. It also demonstrated that RMD strategy was an efficient approach for facilitate screening-out and rapid identification of xenobiotics in biological samples after oral administration of TCMs.

Bakuchiol, the main active component of Psoralea corylifolia, showed a range of significant pharmacological activities, including antimicrobial, antiinflammatory, reduction of bone loss and estrogenic activities. In this research, 12 metabolites, including 11 new compounds, were isolated from the urine and feces of rats after oral administration of bakuchiol, and their structures were elucidated by extensive spectroscopic analysis. The possible metabolic pathways of bakuchiol in rats were proposed, and a rare bile acid conjugation reaction was found. In addition, bakuchiol and its metabolites M1-M3 were studied for their alkaline phosphatase (ALP) activities on MC3T3-E1 cells and cytotoxicity on HKC-8 cells. The data showed that bakuchiol exerted significant effects on ALP activity of MC3T3-E1 cells and cytotoxicity on HKC-8 cells, while M1-M3 only showed ALP activities at 10(-5)M on MC3T3-E1 cells and no cytotoxicity on HKC-8 cells.

A target and nontarget strategy based on in-house chemical components library was developed for rapid and comprehensive analysis of complicated components from traditional Chinese medicine preparation Shuang-Huang-Lian oral liquid. The sample was analyzed by ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry using generic acquisition parameters. Automated detection and data filtering were performed on the UNIFI™ software and the detected peaks were evaluated against an in-house library. As a result, a total of 170 chemical components (110 target compounds and 60 nontarget ones) were identified or tentatively characterized, including 54 flavonoids, 30 phenylethanoid glycosides, 16 iridoid glycosides, 14 lignans, 32 organic acids, 19 triterpenoid saponins and five other types of compounds. Among them, 44 compounds were further confirmed by comparison with reference standards. It was demonstrated that this systematical approach could be successfully applied for rapid identification of multiple compounds in traditional Chinese medicine and its preparations. Furthermore, this work established the foundation for the further investigation on the metabolic fates of multiple ingredients in Shuang-Huang-Lian oral liquid.