Fabrication of amino- and hydroxyl dual-functionalized magnetic microporous organic network for extraction of zearalenone from traditional Chinese medicine prior to the HPLC determination.

Efficient enrichment of trace zearalenone (ZEN) from the complex traditional Chinese medicine (TCM) samples is quite difficult, but of great significance for TCM quality control. Herein, we reported a novel magnetic solid phase extraction (MSPE) strategy for ZEN enrichment using the amino- and hydroxyl dual-functionalized magnetic microporous organic network (Fe3O4@MON-NH2-OH) as an advanced adsorbent combined with the high-performance liquid chromatography (HPLC) determination. Efficient extraction of ZEN was achieved via the possible hydrogen bonding, hydrophobic, and π-π interactions between Fe3O4@MON-NH2-OH and ZEN. The adsorption capacity of Fe3O4@MON-NH2-OH for ZEN was 215.0 mg g-1 at the room temperature, which was much higher than most of the reported adsorbents. Under the optimal condition, the developed Fe3O4@MON-NH2-OH-MSPE-HPLC method exhibited wide linear range (5-2500 µg L-1), low limits of detection (1.4-35 µg L-1), less adsorbent consumption (5 mg), and large enhancement factor (95) for ZEN. The proposed method was successfully applied to detect trace ZEN from 10 kinds of real TCM samples. Conclusively, this work demonstrates the Fe3O4@MON-NH2-OH can effectively extract trace ZEN from the complex TCM matrices, which may open up a new way for the application of MONs in the enrichment and extraction of trace contaminants or active constituents from the complex TCM samples.

Comprehensive metabolome characterization and comparison between two sources of Dragon's blood by integrating liquid chromatography/mass spectrometry and chemometrics.

Dragon's Blood (DB) serves as a precious Chinese medicine facilitating blood circulation and stasis dispersion. Daemonorops draco (D. draco; Qi-Lin-Jie) and Dracaena cochinchinensis (D. cochinchinenesis; Long-Xue-Jie) are two reputable plant sources for preparing DB. This work was designed to comprehensively characterize and compare the metabolome differences between D. draco and D. cochinchinenesis, by integrating liquid chromatography/mass spectrometry and untargeted metabolomics analysis. Offline two-dimensional liquid chromatography/ion mobility-quadrupole time-of-flight mass spectrometry (2D-LC/IM-QTOF-MS), by utilizing a powerful hybrid scan approach, was elaborated for multicomponent characterization. Configuration of an XBridge Amide column and an HSS T3 column in offline mode exhibited high orthogonality (A0 0.80) in separating the complex components in DB. Particularly, the hybrid high-definition MSE-high definition data-dependent acquisition (HDMSE-HDDDA) in both positive and negative ion modes was applied for data acquisition. Streamlined intelligent data processing facilitated by the UNIFI™ (Waters) bioinformatics platform and searching against an in-house chemical library (recording 223 known compounds) enabled efficient structural elucidation. We could characterize 285 components, including 143 from D. draco and 174 from D. cochinchinensis. Holistic comparison of the metabolomes among 21 batches of DB samples by the untargeted metabolomics workflows unveiled 43 significantly differential components. Separately, four and three components were considered as the marker compounds for identifying D. draco and D. cochinchinenesis, respectively. Conclusively, the chemical composition and metabolomic differences of two DB resources were investigated by a dimension-enhanced analytical approach, with the results being beneficial to quality control and the differentiated clinical application of DB.

Multi-level chemical characterization and anti-inflammatory activity evaluation of the polysaccharides from Prunella vulgaris.

Prunella vulgaris (PV) is a medicine and food homologous plant, but its quality evaluation seldom relies on the polysaccharides (PVPs). In this work, we established the multi-level fingerprinting and in vitro anti-inflammatory evaluation approaches to characterize and compare the polysaccharides of P. vulgaris collected from the major production regions in China. PVPs prepared from 22 batches of samples gave the content variation of 5.76-24.524 mg/g, but displayed high similarity in the molecular weight distribution. Hydrolyzed oligosaccharides with degrees of polymerization 2-14 were characterized with different numbers of pentose and hexose by HILIC-MS. The tested 22 batches of oligosaccharides exhibited visible differences in peak abundance, which failed to corelate to their production regions. All the PVPs contained Gal, Xyl, and Ara, as the main monosaccharides. Eleven batches among the tested PVPs showed the significant inhibitory effects on NO production on LPS-induced RAW264.7 cells at 10 µg/mL, but the exerted efficacy did not exhibit correlation with the production regions. Conclusively, we, for the first time, investigated the chemical features of PVPs at three levels, and assessed the chemical and anti-inflammatory variations among the different regions of P. vulgaris samples.

Fingerprinting and characterization of the polysaccharides from Polygonatum odoratum and the in vitro fermented effects on Lactobacillus johnsonii.

Polygonatum odoratum (Yu-Zhu) can be utilized to treat the digestive and respiratory illness. Previous studies have revealed that the underlying therapeutic mechanism of P. odoratum polysaccharides (POPs) is associated with remodeling the gut microbiota. However, POPs in terms of the chemical composition and fermentation activities have been understudied. Here we developed the three-level fingerprinting approaches to characterize the structures of POPs and probed into the beneficial effects on promoting the growth and fermentation of Lactobacillus johnsonii. POPs were prepared by water decoction followed by alcohol sedimentation, while trifluoroacetic acid under different conditions to prepare the hydrolyzed oligosaccharides and monosaccharides. POPs exhibited three main molecular distribution of 601-620 kDa, 4.12-6.09 kDa, and 3.57-6.02 kDa. Hydrolyzed oligosaccharides with degree of polymerization (DP) 2-13 got primarily characterized by analyzing the rich fragmentation information obtained by hydrophilic interaction chromatography/ion mobility-quadrupole time-of-flight mass spectrometry (HILIC/IM-QTOF-MS). Amongst them, the DP5 oligosaccharide was characterized as 1,6,6-kestopentaose. The molecular ratio of Fru: Ara: Glc: Gal: Xyl was 87.72: 0.30: 11.56: 0.19: 0.23. In vitro fermentation demonstrated that 4.5 mg/mL of POPs could significantly promote the growth of L. johnsonii. Co-cultivated with 4.5 mg/mL of POPs, L. johnsonii exhibited stronger antimicrobial activity against Klebsiella pneumoniae. The concentrations of short-chain fatty acids in the POPs-lactobacilli fermented products, including acetic acid, isobutyric acid, and isovaleric acid, were increased. Conclusively, POPs represent the promising prebiotic candidate to facilitate lactobacilli, which is associated with exerting the health benefits.

Polygonatum odoratum polysaccharide attenuates lipopolysaccharide-induced lung injury in mice by regulating gut microbiota.

Polygonatum odoratum is appreciated for its edible and medicinal benefits especially for lung protection. However, the contained active components have been understudied, and further research is required to fully exploit its potential application. We aimed to probe into the beneficial effects of Polygonatum odoratum polysaccharide (POP) in lipopolysaccharide-induced lung inflammatory injury mice. POP treatment could ameliorate the survival rate, pulmonary function, lung pathological lesions, and immune inflammatory response. POP treatment could repair intestinal barrier, and modulate the composition of gut microbiota, especially reducing the abundance of Klebsiella, which were closely associated with the therapeutic effects of POP. Investigation of the underlying anti-inflammatory mechanism showed that POP suppressed the generation of pro-inflammatory molecules in lung by inhibiting iNOS+ M1 macrophages. Collectively, POP is a promising multi-target microecological regulator to prevent and treat the immuno-inflammation and lung injury by modulating gut microbiota.

Steroidal Saponins from Solanum lyratum Thunb.

Four undescribed steroidal compounds along with twenty known compounds were isolated from n-butanol extracted fraction of the whole plants of Solanum lyratum Thunb (SLNF). Their structures were assigned based on analyses of the extensive spectroscopic data (including MS, 1D/2D NMR, and ECD) or comparisons of the NMR data with those reported. Among the knowns, three compounds were isolated from Solanum plants for the first time, while one compound was isolated from S. lyratum for the first time. In addition, the cytotoxicities of these isolates against human colon SW480 and hepatoma Hep3B cells were evaluated by a MTT assay. And, nine of them and SLNF exhibited significant activities against both SW480 and Hep3B cells, while twelve of them significantly inhibited the activities of SW480 cells. This study allows for the exploitation of chemical markers with potential significance in discrimination of Solanum plants, and uncovers the diverse steroidal constituents from S. lyratum dedicated for its future application in cancer treatment.

Multiple heart-cutting two-dimensional liquid chromatography/charged aerosol detector assay of ginsenosides for quality evaluation of ginseng from diverse Chinese patent medicines.

For quality control of Chinese patent medicines (CPMs) containing the same herbal medicine or different herbal medicines that have similar chemical composition, current ″one standard for one species″ research mode leads to poor universality of the analytical approaches unfavorable to discriminate easily confused species. Herein, we were aimed to elaborate a multiple heart-cutting two-dimensional liquid chromatography/charged aerosol detector (MHC-2DLC/CAD) approach to quantitatively assess ginseng from multiple CPMs. Targeting baseline resolution of 16 ginsenosides (noto-R1/Rg1/Re/Rf/Ra2/Rb1/Rc/Ro/Rb2/Rb3/Rd/Rh1/Rg2/Rg3/Rg3(R)/24(R)-p-F11), experiments were conducted to optimize key parameters and validate its performance. A Poroshell 120 EC-C18 column and an XBridge Shield RP18 column were separately utilized in the first-dimensional (1D) and the second-dimensional (2D) chromatography. Eight consecutive cuttings could achieve good separation of 16 ginsenosides within 85 min. The developed MHC-2DLC/CAD method showed good linearity (R2 > 0.999), repeatability (RSD < 6.73%), stability (RSD < 5.63%), inter- and intra-day precision (RSD < 5.57%), recovery (93.76-111.14%), and the limit of detection (LOD) and limit of quantification (LOQ) varied between 0.45-2.37 ng and 0.96-4.71 ng, respectively. We applied it to the content determination of 16 ginsenosides simultaneously from 28 different ginseng-containing CPMs, which unveiled the ginsenoside content difference among the tested CPMs, and gave useful information to discriminate ginseng in the preparation samples, as well. The MHC-2DLC/CAD approach exhibited advantages of high specificity, good separation ability, and relative high analysis efficiency, which also justified the feasibility of our proposed ″Monomethod Characterization of Structure Analogs″ strategy in quality evaluation of diverse CPMs that contained different ginseng.

Characterization and comparison of cardiomyocyte protection activities of non-starch polysaccharides from six ginseng root herbal medicines.

Ginseng is rich of polysaccharides, however, the evidence supporting polysaccharides to distinguish various ginseng species is rarely reported. Focusing on six root ginseng (e.g., Panax ginseng-PG, P. quinquefolius-PQ, P. notoginseng-PN, red ginseng-RG, P. japonicus-PJ, and P. japonicus var. major-PJM), the contained non-starch polysaccharides (NPs) were structurally characterized and compared by both the chemical and biological evaluation. Holistic fingerprinting at three levels (the NPs and the acid hydrolysates involving oligosaccharides and monosaccharides) utilized various chromatography methods, and the treatment of H9c2 cells with the NPs by OGD and H2O2-induced injury models was used to assess the protective effect. NPs from six Panax herbal medicines occupied about 20 % of the total polysaccharides, which were of the highest content in RG and the lowest in PN. NPs from six ginseng exhibited weak differentiations in the molecular weight distribution, while marker oligosaccharides were found to distinguish PN and RG from the others. Glc and GalA were more abundant in the NPs for PG and RG, respectively. NPs from PQ (100/200 µg/mL) showed significant cardiomyocyte protection effect by regulating the mitochondrial functions. This work further testifies the role of polysaccharides in quality control of herbal medicine, with new markers discovered beneficial to distinguish the ginseng.

A practical strategy enabling more reliable identification of ginsenosides from Panax quinquefolius flower by dimension-enhanced liquid chromatography/mass spectrometry and quantitative structure-retention relationship-based retention behavior prediction.

To accurately identify the metabolites is crucial in a number of research fields, and discovery of new compounds from the natural products can benefit the development of new drugs. However, the preferable phytochemistry or liquid chromatography/mass spectrometry approach is time-/labor-extensive or receives unconvincing identifications. Herein, we presented a strategy, by integrating offline two-dimensional liquid chromatography/ion mobility-quadrupole time-of-flight mass spectrometry (2D-LC/IM-QTOF-MS), exclusion list-containing high-definition data-dependent acquisition (HDDDA-EL), and quantitative structure-retention relationship (QSRR) prediction of the retention time (tR), to facilitate the in-depth and more reliable identification of herbal components and thus to discover new compounds more efficiently. Using the saponins in Panax quinquefolius flower (PQF) as a case, high orthogonality (0.79) in separating ginsenosides was enabled by configuring the XBridge Amide and CSH C18 columns. HDDDA-EL could improve the coverage in MS2 acquisition by 2.26 folds compared with HDDDA (2933 VS 1298). Utilizing 106 reference compounds, an accurate QSRR prediction model (R2 = 0.9985 for the training set and R2 = 0.88 for the validation set) was developed based on Gradient Boosting Machine (GBM), by which the predicted tR matching could significantly reduce the isomeric candidates identification for unknown ginsenosides. Isolation and establishment of the structures of two malonylginsenosides by NMR partially verified the practicability of the integral strategy. By these efforts, 421 ginsenosides were identified or tentatively characterized, and 284 thereof were not ever reported from the Panax species. The current strategy is thus powerful in the comprehensive metabolites characterization and rapid discovery of new compounds from the natural products.

Degradation Profiling of Nardosinone at High Temperature and in Simulated Gastric and Intestinal Fluids.

Nardosinone, a predominant bioactive product from Nardostachys jatamansi DC, is well-known for its promising therapeutic applications, such as being used as a drug on anti-inflammatory, antidepressant, cardioprotective, anti-neuroinflammatory, anti-arrhythmic, anti-periodontitis, etc. However, its stability under varying environmental conditions and its degradation products remain unclear. In this study, four main degradation products, including two previously undescribed compounds [2-deoxokanshone M (64.23%) and 2-deoxokanshone L (1.10%)] and two known compounds [desoxo-narchinol A (2.17%) and isonardosinone (3.44%)], were firstly afforded from the refluxed products of nardosinone in boiling water; their structures were identified using an analysis of the extensive NMR and X-ray diffraction data and the simulation and comparison of electronic circular dichroism spectra. Compared with nardosinone, 2-deoxokanshone M exhibited potent vasodilatory activity without any of the significant anti-neuroinflammatory activity that nardosinone contains. Secondly, UPLC-PDA and UHPLC-DAD/Q-TOF MS analyses on the degradation patterns of nardosinone revealed that nardosinone degraded more easily under high temperatures and in simulated gastric fluid compared with the simulated intestinal fluid. A plausible degradation pathway of nardosinone was finally proposed using nardosinonediol as the initial intermediate and involved multiple chemical reactions, including peroxy ring-opening, keto-enol tautomerization, oxidation, isopropyl cleavage, and pinacol rearrangement. Our findings may supply certain guidance and scientific evidence for the quality control and reasonable application of nardosinone-related products.

Comparative Characterization of the Ginsenosides from Six Panax Herbal Extracts and Their In Vitro Rat Gut Microbial Metabolites by Advanced Liquid Chromatography-Mass Spectrometry Approaches.

Ginseng extracts are extensively used as raw materials for food supplements and herbal medicines. This study aimed to characterize ginsenosides obtained from six Panax plant extracts (Panax ginseng, red ginseng, Panax quinquefolius, Panax notoginseng, Panax japonicus, and Panax japonicus var. major) and compared them with their in vitro metabolic profiles mediated by rat intestinal microbiota. Ultrahigh-performance liquid chromatography/ion mobility-quadrupole time-of-flight mass spectrometry (UHPLC/IM-QTOF-MS) with scheduled multiple reaction monitoring (sMRM) quantitation methods were developed to characterize and compare the ginsenoside composition of the different extracts. After in vitro incubation, 248 ginsenosides/metabolites were identified by UHPLC/IM-QTOF-MS in six biotransformed samples. Deglycosylation was determined to be the main metabolic pathway of ginsenosides, and protopanaxadiol-type and oleanolic acid-type saponins were easier to be easily metabolized. Compared with the ginsenosides in plant extracts, those remaining in six biotransformed samples were considerably fewer after biotransformation for 8 h. However, the compositional differences in four subtypes of the ginsenosides among the six Panax plants became more distinct.

[Content determination of ten flavonoids and alkaloids in Gleditsiae Sinensis Fructus, Gleditsiae Fructus Abnormalis, and Gleditsiae Spina].

To study the quality control of three traditional Chinese medicines derived from Gleditsia sinensis [Gleditsiae Sinensis Fructus(GSF), Gleditsiae Fructus Abnormalis(GFA), and Gleditsiae Spina(GS)], this paper established a multiple reaction monitoring(MRM) approach based on ultra-high performance liquid chromatography-triple quadrupole-linear ion-trap mass spectrometry(UHPLC-Q-Trap-MS). Using an ACQUITY UPLC BEH C_(18) column(2.1 mm × 100 mm, 1.7 µm), gradient elution was performed at 40 ℃ with water containing 0.1% formic acid-acetonitrile as the mobile phase running at 0.3 mL·min~(-1), and the separation and content determination of ten chemical constituents(e.g., saikachinoside A, locustoside A, orientin, taxifolin, vitexin, isoquercitrin, luteolin, quercitrin, quercetin, and apigenin) in GSF, GFA, and GS were enabled within 31 min. The established method could quickly and efficiently determine the content of ten chemical constituents in GSF, GFA, and GS. All constituents showed good linearity(r>0.995), and the average recovery rate was 94.09%-110.9%. The results showed that, the content of two alkaloids in GSF(2.03-834.75 µg·g~(-1)) was higher than that in GFA(0.03-10.41 µg·g~(-1)) and GS(0.04-13.66 µg·g~(-1)), while the content of eight flavonoids in GS(0.54-2.38 mg·g~(-1)) was higher than that in GSF(0.08-0.29 mg·g~(-1)) and GFA(0.15-0.32 mg·g~(-1)). These results provide references for the quality control of G. sinensis-derived TCMs.

[Characterization and identification of chemical components in traditional Chinese medicine Psoraleae Fructus based on UHPLC-Q-TOF-MS].

This study was designed to comprehensively characterize and identify the chemical components in traditional Chinese medicine Psoraleae Fructus by establishing an ultra-high performance liquid chromatography/quadrupole time-of-flight mass spectrometry(UHPLC-Q-TOF-MS) method in combination with in-house library. The chromatographic separation conditions(stationary phase, column temperature, mobile phase, and elution gradient) and key MS monitoring parameters(capillary voltage, nozzle voltage, and fragmentor) were sequentially optimized via single-factor experiments. A BEH C_(18) column(2.1 mm×100 mm, 1.7 µm) was finally adopted, with the mobile phase consisting of 0.1% formic acid in water(A) and acetonitrile(B) at the flow rate of 0.4 mL·min~(-1) and column temperature of 30 ℃. Auto MS/MS was utilized for data acquisition in both positive and negative ion modes. By comparison with reference compounds, analysis of the MS~2 fragments, in-house library retrieval and literature research, 83 compounds were identified or tentatively characterized from Psoraleae Fructus, including 58 flavonoids, 11 coumarins, 4 terpenoid phenols, and 10 others. Sixteen of them were identified by comparison with reference compounds, and ten compounds may have not been reported from Psoraleae Fructus. This study achieved a rapid qualitative analysis on the chemical components in Psoraleae Fructus, which provided useful reference for elucidating its material basis and promoting the quality control.

An integrated strategy for comprehensive characterization of metabolites and metabolic profiles of bufadienolides from Venenum Bufonis in rats.

Comprehensive characterization of metabolites and metabolic profiles in plasma has considerable significance in determining the efficacy and safety of traditional Chinese medicine (TCM) in vivo. However, this process is usually hindered by the insufficient characteristic fragments of metabolites, ubiquitous matrix interference, and complicated screening and identification procedures for metabolites. In this study, an effective strategy was established to systematically characterize the metabolites, deduce the metabolic pathways, and describe the metabolic profiles of bufadienolides isolated from Venenum Bufonis in vivo. The strategy was divided into five steps. First, the blank and test plasma samples were injected into an ultra-high performance liquid chromatography/linear trap quadrupole-orbitrap-mass spectrometry (MS) system in the full scan mode continuously five times to screen for valid matrix compounds and metabolites. Second, an extension-mass defect filter model was established to obtain the targeted precursor ions of the list of bufadienolide metabolites, which reduced approximately 39% of the interfering ions. Third, an acquisition model was developed and used to trigger more tandem MS (MS/MS) fragments of precursor ions based on the targeted ion list. The acquisition mode enhanced the acquisition capability by approximately four times than that of the regular data-dependent acquisition mode. Fourth, the acquired data were imported into Compound Discoverer software for identification of metabolites with metabolic network prediction. The main in vivo metabolic pathways of bufadienolides were elucidated. A total of 147 metabolites were characterized, and the main biotransformation reactions of bufadienolides were hydroxylation, dihydroxylation, and isomerization. Finally, the main prototype bufadienolides in plasma at different time points were determined using LC-MS/MS, and the metabolic profiles were clearly identified. This strategy could be widely used to elucidate the metabolic profiles of TCM preparations or Chinese patent medicines in vivo and provide critical data for rational drug use.

A multi-dimensional liquid chromatography/high-resolution mass spectrometry approach combined with computational data processing for the comprehensive characterization of the multicomponents from Cuscuta chinensis.

Challenges encountered in plant metabolites characterization by liquid chromatography/mass spectrometry can arise from the insufficient chromatography separation, the lack of specific database, and low reliability of identification because of the ubiquitous isomerism. Herein, we present an integral approach, by combining comprehensive off-line two-dimensional liquid chromatography/ion mobility-quadrupole time-of-flight mass spectrometry (2D-LC/IM-QTOF-MS), automatic peak annotation, molecular networking, and collision cross section (CCS) prediction, aimed to improve the resolution and reliability in MS-oriented metabolites characterization. Using the seeds of Cuscuta chinensis as a case, the configuration of an XBridge Amide column of hydrophilic interaction chromatography (HILIC) and a Zorbax SB-Aq column of reversed-phase chromatography (RPC), in an off-line mode, showed the orthogonality of 0.73 and effective peak capacity of 4361. Data-independent high-definition MSE (HDMSE) in the negative mode could enable high-coverage MS2 data acquisition and enhance the ions resolution, while computational peak annotation workflows facilitated by UNIFITM and Global Natural Products Social Molecular Network (GNPS) could efficiently characterize the targeted and untargeted compound analogs. A total of 302 compounds were identified or tentatively characterized, and 109 thereof were unreported. Moreover, CCS prediction (www.allccs.zhulab.cn) provided more possibilities to distinguish 12 pairs of isomers in the lack of reference standards. The 2D-LC/IM-QTOF-MS approach enabled the collection of five dimension of data related to each component (tR by HILIC and RPC, CCS, m/z in MS1 and MS2), and the intelligent metabolites characterization with more reliable MS data. Conclusively, the established integral strategy can be utilized in metabolome analysis to support the quality control of herbal medicines.

An ion mobility-enabled and high-efficiency hybrid scan approach in combination with ultra-high performance liquid chromatography enabling the comprehensive characterization of the multicomponents from Carthamus tinctorius.

Liquid chromatography/mass spectrometry (LC/MS) is extensively applied for the untargeted/targeted analyses of the herbal components, utilizing data-dependent acquisition (DDA) or data-independent acquisition (DIA) to record the fragmentation information useful for the structural elucidation. A new trend recently has emerged by integrating DDA and DIA to render the hybrid scan, which, unfortunately, has rarely been reported. Herein, by using the Vion™ ion-mobility quadrupole time-of-flight mass spectrometer, a hybrid scan strategy (HDMSE-HDDDA) was presented and validated by the untargeted characterization of the multicomponents from Carthamus tinctorius (safflower), in combination with reversed-phase ultra-high performance liquid chromatography (RP-UHPLC). Good chromatographic separation was achieved on an HSS T3 column within 26 min, while HDMSE-MS/MS was used to acquire the collision-induced dissociation MS2 data in the negative mode. Automatic workflows (e.g., data correction, precursors/product ions matching, and peak annotation) were well established on UNIFI™ (incorporating an in-house library recording 261 known compounds) to process the obtained MS2 data. Compared with single DDA or DIA, the hybrid approach of HDMSE-HDDDA better balanced between the coverage and reliability, led to high-definition MS spectra, offered useful collision cross section (CCS) information, and showed satisfactory identification performance comparable to MSE. A total of 141 components (involving 41 quinochalcones, 66 flavanols/flavones, 11 flavanones, 6 organic acids, 1 polyacetylene, and 16 others) were characterized from safflower. Moreover, CCS prediction could assist isomers characterization, to some extent. Conclusively, this hybrid scan approach enables a dimension-enhanced MS data acquisition strategy providing the complementary structural information, which more suits the chemical characterization of complex samples.

Advances and challenges in ginseng research from 2011 to 2020: the phytochemistry, quality control, metabolism, and biosynthesis.

Covering: 2011 to the end of 2020Panax species (Araliaceae), particularly P. ginseng, P. quinquefolius, and P. notoginseng, have a long history of medicinal use because of their remarkable tonifying effects, and currently serve as crucial sources for various healthcare products, functional foods, and cosmetics, aside from their vast clinical preparations. The huge market demand on a global scale prompts the continuous prosperity in ginseng research concerning the discovery of new compounds, precise quality control, ADME (absorption/disposition/metabolism/excretion), and biosynthesis pathways. Benefitting from the ongoing rapid development of analytical technologies, e.g. multi-dimensional chromatography (MDC), personalized mass spectrometry (MS) scan strategies, and multi-omics, highly recognized progress has been made in driving ginseng analysis towards "systematicness, integrity, personalization, and intelligentization". Herein, we review the advances in the phytochemistry, quality control, metabolism, and biosynthesis pathway of ginseng over the past decade (2011-2020), with 410 citations. Emphasis is placed on the introduction of new compounds isolated (saponins and polysaccharides), and the emerging novel analytical technologies and analytical strategies that favor ginseng's authentic use and global consumption. Perspectives on the challenges and future trends in ginseng analysis are also presented.

A novel hybrid scan approach enabling the ion-mobility separation and the alternate data-dependent and data-independent acquisitions (HDDIDDA): Its combination with off-line two-dimensional liquid chromatography for comprehensively characterizing the multicomponents from Compound Danshen Dripping Pill.

Data-dependent acquisition (DDA) and data-independent acquisition (DIA)-based MSn strategies are extensively applied in metabolites characterization. DDA gives accurate MSn information, but receives low coverage, while DIA covers the entire mass range, but the precursor-product ions matching often yields false positives. Currently available MS scan approaches rarely integrate DIA and DDA within a duty circle. Utilizing a Vion™ IM-QTOF (ion mobility-quadrupole time-of-flight) mass spectrometer, we report a novel hybrid scan approach, namely HDDIDDA, which involves three scan events: 1) IM-enabled full scan (MS1), 2) high-definition MSE (HDMSE) of all precursor ions (MS2); and 3) high-definition DDA (HDDDA) of top N precursors (MS2). As a proof-of-concept, the HDDIDDA approach combined with off-line two-dimensional liquid chromatography (2D-LC) was applied to characterize the multiple ingredients from a reputable Chinese patent medicine, Compound Danshen Dripping Pill (CDDP) used for treating the cardiovascular diseases. An off-line 2D-LC system by configuring an XBridge Amide column and an HSS T3 column showed a measurable orthogonality of 0.92 and enhanced the separation of co-eluting components. A fit-for-purpose HDDIDDA methodology was developed in the negative mode to characterize saponins and salvianolic acids, while tanshinones in the positive mode. Computational workflows to efficiently process the acquired HDMSE and HDDDA data were established, and the searching of an in-house CDDP library (recording 712 compounds) eventually characterized 403 components from CDDP, indicating approximate 12-fold improvement compared with the previous report. The HDDIDDA approach can measure collision cross section of each component, and merges the merits of DIA and DDA in MS2 data acquisition.

Treponema pallidum (Syphilis) Antigen TpF1 Induces Activation of Macrophages and Accelerates P2X7R-Induced NLRP3-Dependent Release of IL-1ß.

BACKGROUND: Syphilis is a chronic infectious disease caused by Treponema pallidum (Tp) infection, which causes local inflammation in the host. TpF1 is an oligomeric protein expressed by the Tp-infected host that can induce the host immune response. There are few studies regarding the role of TpF1 in macrophage activation and the subsequent release of cytokines. OBJECTIVE: The objective of this study is to elucidate the effects of TpF1 on the pathological process of Syphilis. In addition, we explored how purinergic 2X7 (P2X7R) induced NOD-like receptor family protein 3 (NLRP3) -dependent release of interleukin-1ß (IL-1ß) and the underlying mechanisms. METHODS: We explored the influence of TpF1 on cytokine release by macrophages using qRT-PCR and ELISA. The specific phenotype of activated macrophages was determined by flow cytometry. RESULTS: TpF1 was able to activate macrophages and induce the M1 macrophage phenotype. Moreover, TpF1 activated the NLRP3 inflammasome in macrophages, which was mediated by P2X7R. CONCLUSION: The Tp-induced protein TpF1 is able to induce macrophage activation and P2X7R-induced NLRP3-dependent release of IL-1ß. Our findings provide a theoretical basis for clarifying the clinical symptoms and pathogenesis of syphilis.

Multi-level fingerprinting and cardiomyocyte protection evaluation for comparing polysaccharides from six Panax herbal medicines.

The role of polysaccharides in quality control of ginseng is underestimated. Large-scale comparison on the polysaccharides of Panax ginseng (PG), P. quinquefolius (PQ), P. notoginseng (PN), Red ginseng (RG), P. japonicus (ZJS), and P. japonicus var. major (ZZS), was performed by both chemical and biological approaches. Holistic fingerprinting at polysaccharide and the hydrolyzed oligosaccharide and monosaccharide levels utilized various chromatography methods, while OGD and OGD/R models on H9c2 cells were introduced to evaluate the protective effects on cell viability and mitochondrial function. Polysaccharides from six ginseng species exhibited remarkable content difference (RG > PG/ZZS/ZJS/PQ > PN), but weak differentiations in molecular weight distribution and oligosaccharide profiles, while Glc and GalA were richer for monosaccharide compositions of PG and RG polysaccharides, respectively. RG polysaccharides (25/50/100 µg/mL) showed significant cardiomyocyte protection by regulating mitochondrial functions. These new evidences may provide support for the supplementary role of polysaccharides in quality control of ginseng.