Identifying the structures of metabolites is of great importance towards elucidating the therapeutic material basis of traditional Chinese medicines （TCMs）. However， current studies lack sufficient reference standards， hindering the structural identification of metabolites. This study establishes a strategy guided by the retention times and mass spectrometric information of microsomal metabolites. By coupling liquid chromatography-mass spectrometry （LC-MS） to trace and isolate target compounds from the raw materials， these compounds served as reference standards to confirm the structures of microsomal metabolites， thereby elucidating the metabolic pathways. Peucedani Radix （PR， Chinese name： Qianhu） consists of the dried roots of Peucedanum praeruptorum Dunn and contains abundant angular-type pyranocoumarins （APs）， which serve as primary chemical constituents and effective compound clusters as well. From the structural standpoint， C-3′ and C-4′ sites of APs are frequently substituted by acetoxyl， angeloxyl， isovaleroxyl and some other moieties， leading to the extensive occurrences of regio-isomers， which not only possess comparable structures but also similar chemical properties. Previous studies have preliminarily unveiled that APs primarily undergo hydrolysis and acyl migration in vivo， and these hydrolytic metabolites can be observed in both in vitro metabolism and the raw materials. In this study， a pair of regio-isomers from PR， i.e.， （+）-（3′S，4′S）-praeruptorin A （（+）-PA） and （+）-（3′R，4′R）-pteryxin （（+）-Pte）， was incubated separately with human liver microsomes （HLMs） in the presence of necessary cofactors to simulate the metabolic pathways occurring in vivo. After carefully processing LC-MS datasets and applying the well-defined mass fragmentation rules of APs， eight metabolites were detected and putatively identified for （+）-PA， including three hydrolysis products together with five oxidation products， whereas nine metabolites were observed and putatively identified for （+）-Pte， encompassing five hydrolysis products and four oxidation products. Noteworthily， phase I metabolism of （+）-Pte in HLMs was profiled for the first time. Various column chromatographic tools， such as silica gel column chromatography， C₁₈ reversed-phase chromatography （ODS）， Sephadex LH-20 and semi-preparative high performance liquid chromatography equipped with achiral or chiral columns， were employed to isolate compounds-of-interest from PR extracts under the guidance of LC-MS through taking both retention time and MS/MS information into account. A total of nine AP compounds were successfully purified from PR extracts， and notably， four pairs of AP enantiomers were involved. By matching ¹H- and ¹³C-NMR spectral information with the data archived in the literature， these compounds were identified as （3′S，4′S）-qianhucoumarin C （1a）， （3′R，4′R）-qianhucoumarin C （1b）， （3′S，4′S）-qianhucoumarin B （2a）， （3′R，4′R）-qianhucoumarin B （2b）， （3′S，4′S）-3′-angeloylkhellactone （3a）， （3′R，4′R）-3′-angeloylkhellactone （3b）， （3′S，4′S）-4′-angeloylkhellactone （4a）， （3′R，4′R）-4′-angeloylkhellactone （4b） and （3′S，4′S）-isoepoxypteryxin （5）. Thereof， compounds 1b and 2b represent new configurational structures of known planar structures. The study summarized mass fragmentation rules for APs and achieved accurate identification of two pairs of regio-isomers： qianhucoumarin B and C， and 3′- and 4′-angeloylkhellactone. Thereafter， in vitro metabolites of （+）-PA and （+）-Ptewere successfully identified by comparing the retention times and MS/MS information with authentic compounds， including cis-khellactone， qianhucoumarin B， qianhucoumarin C， 3′-angeloylkhellactone， 4′-angeloylkhellactone and （3′S，4′S）-isoepoxypteryxin. Regarding the three hydrolytic metabolites， i.e.， cis-khellactone， qianhucoumarin B and qianhucoumarin C， they were shared by （+）-PAand （+）-Pte. Hydrolysis and oxidation reactions， as well as acyl migration， served as the primary metabolic pathways for APs in HLMs. In this study， （+）-PAand （+）-Ptewere selected as representatives to demonstrate that it is feasible to obtain their metabolites from PR extracts through LC-MS-guided isolation and purification. After structural identification through analyzing ¹H- and ¹³C-NMR spectral information， these pure compounds were eligible authentic compounds enabling confirmative structural identification of metabolites， providing pronounced insights into the primary metabolic pathways of APs and offering an effective approach for the accurate identification of metabolite structures through isolating authentic compounds that are available in the original TCM. However， due to differences in metabolism between in vitro and in vivo， the study failed to construct a definitive structure-activity relationship for the metabolites. Future studies should be devoted to validating these findings through performing in vivo investigations.

Exosomes are nanometer-sized secretory vesicles that play a pivotal role in mediating intercellular communication. Recently， exosomes derived from herbal medicine have garnered significant attention owing to their unique composition features and therapeutic potential. Glycyrrhiza uralensis， a traditional Chinese medicinal and edible plant， is rich in flavonoids， terpenoids， polyketides， and phenolic acids. However， the long-term consequences of overharvesting and artificial cultivation have led to inconsistent quality of licorice materials. Plant tissue culture technology offers a sustainable approach to preserve medicinal plant resources， enabling year-round production independent of seasonal and environmental constraints. To address these challenges and explore the therapeutic potential of herb-derived exosomes， this study employed a laboratory-constructed temporary immersion bioreactor system （TIBS） for G. uralensis cultivation and exosomes isolation. The key experimental procedures are summarized as follows： sterile buds derived from G. uralensis seeds were used as explants. The callus proliferation medium consisted of Murashige and Skoog medium （MSM） supplemented with 1.18 mg/L 6-benzylaminopurine （6-BA）， 1.44 mg/L naphthaleneacetic acid （NAA） and 1.44 mg/L 2，4-dichlorophenoxyacetic acid （2，4-D）， yielding a maximum proliferation coefficient of 10.81 after 21 days. For shoot proliferation， sterile plantlets were generated on MSM medium supplemented with 1 mg/L 6-BA， 0.05 mg/L NAA， and 150 mL/L coconut milk （CM）， which produced sterile plantlets within 28 days of culture. When cultured in the TIBS system with an immersion frequency of 5 min every 6 h， the callus proliferation coefficient reached 13.12 after 28 days of culture. Exosomes were successfully isolated from the TIBS-cultured G. uralensis calli using differential centrifugation and ultracentrifugation protocols. Comprehensive characterization was performed to validate the identity， purity， and stability of isolated exosomes. Morphological observation via transmission electron microscopy （TEM） revealed typical cup-shaped vesicles with a distinct lipid bilayer structure， which is consistent with the canonical exosomes morphology. Dynamic light scattering （DLS） analysis determined an average hydrodynamic diameter of 56 nm. The exosomes preparation exhibited a particle concentration of 1.07×10¹² particles/mL， and a zeta potential of （-15.030±3.815） mV， indicating excellent colloidal stability suitable for biological applications. To decipher the molecular cargo of these exosomes， untargeted metabolomic profiling was conducted. A total of 1 760 metabolites were identified across callus and exosomes samples， of which 1 068 differential metabolites （DMs） distinguished exosomes from parental calli. Notably， 195 DMs were significantly enriched in exosomes. KEGG （Kyoto Encyclopedia of Genes and Genomes） pathway enrichment analysis revealed enrichment in 11 metabolic pathways， primarily involving terpenoids， flavonoids， alkaloids， fatty acids， and phenylpropanoids. Bioactive compounds of interest included azelaic acid， phloretin， and senkyunolide C， highlighting their potential pharmacological significance. In summary， this study demonstrates that the TIBS-based in vitro platform enables efficient， scalable， and consistent production of G. uralensis-derived exosomes with homogeneous morphology and specialized metabolite profiles. The successful isolation， comprehensive characterization， and identification of bioactive metabolite enrichment establish a robust foundation for mechanistic investigations and translational applications. These exosomes exhibited promising therapeutic potential in anti-inflammatory， antioxidant， antitumor， and antibacterial biomedicine. Furthermore， this bioreactor system addresses the quality inconsistency issue in natural G. uralensis resources， paving new ways for developing innovative herbal exosome-based nanomedicines.

The composition of natural products is usually very complex. During the separation of natural products， high loading capacity and satisfactory chromatographic resolution are often hardly achieved. To address this limitation， various modifiers were added in the mobile phase to alter the chromatographic behavior. Here， deep eutectic solvents （DESs） were chosen to accomplish this objective on C18 column. Changes in retention factor， overloaded peak shape and adsorption capacity were systematically investigated and interpreted. The application potential of DESs in the separation of natural products was then evaluated. For ionizable compounds， the addition of DESs could compress the peak width and enhance the resolution. This is closely related to the electrostatic interactions provided by hydrogen bond acceptor （HBA） of DESs. For ionizable basic compounds like nortriptyline hydrochloride， they could remain in a cationic state constantly. Consequently， ion-exchange interactions occurred between nortriptyline cation and residual silanol anion， which resulted in severe peak tailing. However， this interaction could be shielded by the addition of HBA of DESs （such as choline chloride （ChCl））. As the pH of the mobile phase increased， the dissociation of silanol groups was enhanced. This rendered the shielding effect more prominent and led to a higher peak compression of nortriptyline hydrochloride. Notably， although the mobile phase was acidic， the dissociation of silanol groups cannot be completely suppressed. It means that this modulating effect of HBA never disappears. Based on these principles， the semi-preparative separation of berberine from Coptis chinensis extracts was successfully improved. The improvement in the separation of acidic compounds by DESs depends on the ion-pair formation. This effect is highly correlated with the mobile phase pH and salt concentration. Here， benzoic acid was chosen as a model to investigate the overload behavior and adsorption. The results indicated that under neutral conditions， ChCl addition converted the sigmoidal adsorption to a linear type. It can be attributed to the ion-pair formation between benzoate anions and HBA cations. Mutual repulsion between benzoate anions was therefore reduced and adsorption capacity correspondingly enhanced. Furthermore， the results of the overload experiment indicated that ChCl can effectively weaken the solute-solute interactions. It is worth noting that the mobile phase pH is barely influenced after the addition of ChCl. Remarkably， these features are of critical importance for the optimization of preparative chromatographic methods. The beneficial effect of HBAs was validated in the semi-preparative separation of chlorogenic acid from Lonicera japonica extracts. For neutral compounds like morroniside， the HBA in DESs cannot affect their chromatographic behavior through electrostatic interactions. However， there are usually many ionizable impurities surrounding the neutral compounds in natural products. Their chromatographic behavior can be modulated by the HBAs in DESs. The separation of neutral compounds in natural products could be indirectly optimized by modulating their chromatographic behavior. This view was also confirmed by the separation of morroniside from Cornus officinalis extracts.

Earthworms are a commonly used animal-derived traditional Chinese medicine in China， with proteins and polypeptides as their main active ingredients. However， the existing methods for separating and detecting earthworm polypeptides have problems such as insufficient preparation capacity and poor separation reproducibility. To address these challenges， this study used the iron porphyrin metal-organic framework material PCN-222（Fe） to specifically adsorb the earthworm polypeptide component L-PeakⅡ， with an adsorption capacity of 139 mg/g. First of all， PCN-222（Fe） material was successfully prepared via the one-pot method. The earthworm protease hydrolysate was obtained through alkali extraction and acid precipitation， and trypsin enzymatic hydrolysis. Then， PCN-222（Fe） was further used as a solid-phase adsorbent and combined with liquid chromatography to rapidly separate and prepare the polypeptide component L-PeakⅡ from complex earthworm enzymatic hydrolysate of the Chinese medicine. Qualitative analysis was performed by HPLC， and quantitative analysis was conducted with a BCA assay kit. HPLC analysis of the earthworm protease hydrolysate revealed multiple chromatographic peaks. After enrichment with PCN‑222（Fe） and subsequent elution， the eluted sample exhibited fewer peaks， with a well‑defined peak appearing at a retention time of approximately 15 min. In order to obtain the optimal experimental conditions for the enrichment of earthworm polypeptides， influencing factors including enzyme hydrolysate mass concentration， pH value， elution solvent type， and elution time were investigated and optimized. The results showed that optimal enrichment was achieved by incubating at an enzymatic hydrolysate mass concentration of 3.0 mg/mL and pH of 9， followed by elution with 50% acetonitrile containing 0.2% trifluoroacetic acid for 2 h. Finally， the earthworm-derived polypeptide component was obtained using a high-pressure preparative chromatographic system， achieving a purity exceeding 95%. The integrated approach employed in this study， which combines PCN-222（Fe） material with preparative chromatography， enables the efficient preparation of the target polypeptide L-PeakⅡ from medicinal earthworm samples. This strategy provides a valuable reference for the separation and purification of polypeptide components from complex animal-derived traditional Chinese medicines. Moreover， it offers a more robust， scalable， and reproducible pathway for harnessing the therapeutic potential of natural product peptides， thereby bridging the gap between traditional medicine and modern pharmaceutical analysis.

Ginger， the fresh rhizome of Zingiber officinale Rosc. from the Zingiberaceae family， is highly valued worldwide for its unique aromatic profile and distinct pungent flavor. It is widely utilized both as a culinary spice and a natural food additive. Globally， ginger comprises 53 known genes and approximately 1 300 varieties， predominantly cultivated in tropical and subtropical regions. Major producing countries include India， China， Indonesia， and Nigeria， with China ranking as the second-largest producer. In many Asian countries， such as China， Korea， and Japan， ginger has long been recognized not only for its culinary applications but also for its medicinal properties. It is traditionally employed to alleviate cold symptoms， promote body warmth， reduce nausea， suppress coughs by clearing phlegm， and even neutralize toxins from seafood consumption. The efficacy and safety of ginger as a medicinal herb are largely determined by the content and purity of its active constituents， such as gingerols and shogaols. Thus， accurate quality assessment is essential to ensure its therapeutic value. Moreover， the contents of these bioactive compounds can vary considerably depending on the geographical origin. In China， high-quality medicinal ginger is primarily cultivated in provinces such as Yunnan， Sichuan， Guizhou and Henan. Given the complexity and diversity of ginger’s chemical composition， it is crucial to understand its characteristic components and their variation with origin. This knowledge supports not only accurate quality evaluation but also reliable traceability of ginger sources. Ultimately， it facilitates the selection of ginger with superior medicinal properties for direct use in traditional medicine or as high-quality raw material in pharmaceutical development. However， conventional analytical techniques for determining origin and quality， such as high performance liquid chromatography （HPLC） and gas chromatography-mass spectrometry （GC-MS）， are often time-consuming， require extensive sample preparation， and consume large volumes of solvents. To overcome these limitations， this study developed a high-throughput method based on internal extractive electrospray ionization mass spectrometry （iEESI-MS） for rapid compositional profiling and origin authentication of ginger. Samples were collected from four major ginger-producing regions in China： Yunnan， Sichuan， Guizhou， and Henan. Each sample was minimally processed into fragments， with only 5.0 mg used for analysis. The fragments were placed on a filter membrane inside a custom-built iEESI-MS device. Key instrumental parameters such as the extraction solvent composition， ion transfer tube temperature， spray voltage， and solvent flow rate were systematically optimized to enhance detection sensitivity and reproducibility. Under optimal conditions， real-time extraction and ionization of chemical constituents from ginger tissue were achieved， yielding representative mass spectral fingerprints for each geographical origin. Multivariate statistical tools were applied to interpret the complex mass spectrometry data. Principal component analysis （PCA） provided an overview of sample distribution and revealed inherent clustering trends according to origin. Partial least squares discriminant analysis （PLS-DA） further improved the classification accuracy by filtering out unrelated variations and emphasizing the ions that contributed most to inter-regional differences. Through this approach， 27 compounds were consistently detected and identified， among which three key markers， such as 6-gingerol， 8-gingerol， and L-serine， were selected as characteristic of origin-related variation. A quantitative method was developed for these three markers， demonstrating excellent linearity across a broad content range （2.0-20 000.0 μg/g） with coefficients of determination （R²） no less than 0.996. Sensitivity was assessed in terms of limits of detection and limits of quantification， which ranged from 3.0 μg/g to 20.0 μg/g and 10.0 μg/g to 50.0 μg/g， respectively. Recoveries varied between 98.8% and 100.9%， indicating high accuracy， while repeatability was excellent， with relative standard deviations （RSDs） no more than 1.6%. When the established method was applied to ginger samples from different regions， heatmap visualization clearly illustrated the correlation between geographic origin and the abundance of the three characteristic compounds. For instance， Yunnan samples exhibited notably higher contents of certain gingerols， whereas those from Henan were richer in L-serine. This chemical fingerprinting strategy offers a reliable and efficient means for rapid origin verification and quality assessment. In conclusion， the iEESI-MS platform developed in this study combines minimal sample preparation， rapid analysis， and high sensitivity， making it well-suited for high-throughput applications. It holds significant potential not only for ginger authentication but also for quality control of other medicinal plants， food traceability systems， and the protection of geographically indicated products. Future studies may expand the database to include more regions and varieties， further improving the robustness and general applicability of the model.

Antarctic krill has the largest biomass of any wild animal species on Earth. Krill oil extracted from Antarctic krill is rich in phospholipids， and is a major natural source of marine phospholipids. In marine phospholipids， eicosapentaenoic acid （EPA） and docosahexaenoic acid （DHA） are esterified at the sn-2 position of glycerol. This unique structural configuration endows them with the dual physiological functions of both phospholipids and Ω-3 polyunsaturated fatty acids. Compared with terrestrial phospholipids， marine phospholipids exhibit superior bioactivity in several key aspects， including neurotrophic support， anti-atherosclerotic effects， and lipid metabolism regulation， thereby conferring significant application value in pharmaceuticals， health foods， and functional foods. However， pharmaceutical excipient applications require phospholipid purity above 90%， a standard also required for high-end pharmaceutical formulations and active pharmaceutical ingredients. Currently， commercial krill oil products vary significantly in quality. Their total phospholipid content ranges from only 30% to 60%. Therefore， purifying phospholipids from Antarctic krill is essential. This process holds practical value for improving the quality standards of marine phospholipids. Hydrophilic interaction liquid chromatography （HILIC） separates polar compounds through multiple mechanisms， relying on polar interactions between the hydrophilic headgroups of phospholipids and the stationary phase， as well as partitioning within the water-rich layer on the stationary phase surface. In this study， a HILIC-based method was developed for the purification of phospholipids from Antarctic krill oil. Industrial-scale preparative chromatography requires stable stationary phases. Three HILIC-compatible stationary phases were systematically evaluated. These were unmodified bare silica gel， Diol （3% carbon）， and Diol-H （7% carbon）. Diol showed favorable retention and resolution. Therefore， Diol was selected as the optimal stationary phase. The superior performance of the Diol phase compared to Diol-H suggests that a moderate carbon content is crucial. Excessive hydrophobicity in Diol-H likely interfered with the specific hydrogen bonding interactions required for separating polar lipid headgroups. Ethanol and water were chosen as the mobile phases， because these solvents have low toxicity. Ethanol-water proportions were investigated to develop a three-step elution program： washing with 100% ethanol， elution with 95% ethanol， and column re-equilibration with 80% ethanol. Based on chromatographic plate and rate theories， a Diol column （250 mm×4.6 mm） was used. Isocratic elution with 95% ethanol was applied. The relationship between flow rate， theoretical plate number， and peak width was evaluated. An optimal flow rate was selected. Peak broadening under high sample loading was examined. This determined the maximum loading capacity. It was optimized at 8% of the stationary phase mass. Separation remained efficient at this level. Resolution did not decrease significantly. The final purification protocol involved washing with 2 column volumes of 100% ethanol， eluting the target phospholipid fraction with 5.5 column volumes of 95% ethanol， and re-equilibrating the column with 2.5 column volumes of 80% ethanol. Finally， scale-up separation was performed using a DAC50 column. The crude oil contained 57.88% phospholipids. Four fractions were collected based on online chromatograms. Each fraction was concentrated， dried， and then analyzed. The recovery of the purified product was 49.15%. Purity was measured using the molybdenum blue colorimetric method （GB/T 5537-2008）. Purity reached 95.42%. The overall phospholipid recovery was 81%. EPA and DHA were quantified via external standard method （GB 5009.168-2016）. EPA and DHA contents were 18.5% and 7.18%， respectively. This method efficiently prepares pharmaceutical-grade phospholipids. The process is environmentally friendly and easily scalable for industrial manufacturing. This approach supports the purification of marine phospholipids from Antarctic krill oil and the high-purity concentrate serves as a reference material. It supports clinical research on krill-derived lipids and promotes applications in drug delivery and nutraceuticals. This work improves quality standards for marine lipid products and contributes to the sustainable use of Antarctic krill resources. Preparative liquid chromatography is a mature technology widely used for separating complex samples like drugs and natural products. This study investigated HILIC for purifying phospholipids from Antarctic krill oil. The research provides a technical basis for selecting preparation methods.

New pollutants are recently identified or recognized chemical substances. They pose risks to ecosystems or human health. Many are not yet regulated or lack effective control measures. These pollutants show biological toxicity， environmental persistence， and bioaccumulation. They threaten human health. In recent years， their detection frequency in food has increased. Efficient detection technologies are urgently needed. Sample pretreatment is key for analyzing new pollutants in food. The core of pretreatment lies in the preparation and selection of adsorbent materials. Covalent organic frameworks （COFs） are porous crystalline materials. They are formed by light elements linked through covalent bonds. COFs have highly ordered crystal structures. Their pore sizes can be adjusted. Surface properties are functionalizable. They show excellent chemical and thermal stability. Composite covalent organic framework materials combine COFs with other materials. This is achieved through physical or chemical methods. The composites exhibit synergistic effects. They retain the unique properties of both COFs and the other materials. This article reviews common sample pretreatment techniques for new pollutants in food. These include solid-phase extraction （SPE）， solid-phase microextraction （SPME）， stir bar sorptive extraction （SBSE）， dispersive solid-phase extraction （DSPE）， and magnetic solid-phase extraction （MSPE）. SPE is a chromatographic technique. It removes impurities from solid or liquid samples. It also enriches target compounds. SPE offers high enrichment factors and low solvent use. It is easy to automate. SPME balances samples between solid and liquid phases. It integrates sampling， extraction， and concentration. SPME uses little or no solvent. It is simple and automatable. It can be coupled with other techniques online. SBSE evolved from SPME. It has a larger stationary phase volume and higher capacity. SBSE uses a stir bar with a magnetic core. The bar is coated with an extraction layer. Stirring ensures full contact with analytes. SBSE is solvent-free or uses minimal solvent. It is accurate， fast， and easy to automate. DSPE disperses adsorbents into sample matrices. It increases contact area between adsorbents and analytes. DSPE simplifies sample processing. It avoids sample loss. MSPE uses magnetic or magnetizable materials as adsorbents. It captures target analytes efficiently. MSPE is simple to prepare and separate. Pipette-tip-SPE （PT-SPE） is a newer technique. It packs adsorbents into pipette tips. PT-SPE is flexible， low-cost， and needs small sample volumes. This article details types of composite COF materials. These include magnetic COF （MCOF）， sponge-COF， molecularly imprinted polymer-COF （MIP-COF）， metal-organic framework-COF （MOF-COF）， and electrospun-COF. MCOF combines COFs with magnetic nanoparticles. It enables quick separation under a magnetic field. Sponge-COF grows COFs on sponge fibers. It enhances adsorption capacity and mass transfer. MIP-COF integrates molecularly imprinted polymers with COFs. It offers specific recognition sites. MOF-COF combines metal-organic frameworks with COFs. It introduces metal active sites. Electrospun-COF embeds COFs into polymer nanofibers. It improves mechanical performance and stability. These composite materials are utilized in the pretreatment of food samples， where they effectively enrich trace levels of new pollutants， including perfluoroalkyl substances， antibiotics， personal care products， endocrine disruptors， and flame retardants， within complex food matrices. When coupled with analytical techniques such as high-performance liquid chromatography （HPLC）， HPLC-tandem mass spectrometry （HPLC-MS/MS）， gas chromatography （GC）， gas chromatography-mass spectrometry （GC-MS）， molecular fluorescence spectroscopy， and Raman spectroscopy， they facilitate highly accurate and sensitive detection of these new pollutants in food products. Future directions include improving synthesis methods. Current methods are time-consuming and costly. New techniques like water-phase synthesis are promising. Multifunctional composites are needed. They should adsorb multiple pollutant types. Automated and high-throughput extraction technologies will be developed. Green and scalable production processes are essential for industrial applications. In conclusion， composite COF materials show great potential. They enhance the efficiency and accuracy of food pollutant analysis. Further research will expand their applications and improve performance.

This study employed an integrated strategy combining ultra performance liquid chromatography-linear trap quadrupole-Orbitrap mass spectrometry （UPLC-LTQ-Orbitrap-MS）， network pharmacology， molecular docking simulations， and in vivo animal experimentation to systematically analyze the active components of Synotissolidaginea （S. solidaginea） and to elucidate its underlying therapeutic mechanism in the treatment of acute eczema. Initially， the UPLC-LTQ-Orbitrap-MS method was utilized to perform a comprehensive chemical profiling of S. solidaginea extract. Based on accurate mass， multistage fragmentation spectra， and cross-validation with literature and reference standards， 59 constituents were unambiguously or tentatively identified. Subsequently， a network pharmacology framework was established to predict the potential pharmacological mechanisms of the identified compounds. Among the characterized constituents， seven bioactive ingredients—namely isorhamnetin， kaempferol， luteolin， quercetin， myricetin， betaine， and rosmarinic acid—were selected as the primary active components based on their relative abundance and favorable oral bioavailability and drug-likeness properties as predicted by the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform （TCMSP）. The corresponding potential protein targets of these seven compounds were systematically screened using the TCMSP database and the SwissTargetPrediction tool， resulting in the identification of 192 component-associated targets after the removal of duplicate entries. Concurrently， a comprehensive collection of disease-associated targets was curated for acute eczema. By querying the GeneCards database and the Online Mendelian Inheritance in Man （OMIM） database with the keyword “acute eczema”， a total of 2 214 distinct therapeutic targets relevant to the pathogenesis and progression of the dermatological condition were retrieved. Through the construction of a Venn diagram to map the component-related targets onto the disease-specific target set， a core network comprising 75 overlapping targets was identified as the potential therapeutic intersection through which S. solidaginea exerts its pharmacological effects against acute eczema. To further investigate the functional interactions and biological significance of these 75 intersecting proteins， a protein-protein interaction （PPI） network was constructed using the STRING database （version 12.0） and subsequently visualized and topologically analyzed using Cytoscape 3.10.1 software. Network topological analysis， specifically evaluating the degree centrality， betweenness centrality， and closeness centrality， highlighted SRC proto-oncogene tyrosine-protein kinase and matrix metalloproteinase 9 （MMP9） as the most prominent hub nodes within the network， suggesting their pivotal roles in mediating the therapeutic activity. Furthermore， Gene Ontology （GO） functional annotation and Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway enrichment analyses were conducted on the 75 potential targets. The GO enrichment analysis indicated that these targets were predominantly involved in biological processes related to the regulation of inflammatory responses， cellular response to oxidative stress， and positive modulation of cytokine production. The KEGG pathway analysis further revealed that the therapeutic effects of S. solidaginea are likely orchestrated through the modulation of several critical signaling cascades， with the PI3K/Akt signaling pathway being the most significantly enriched pathway. To validate the credibility of these computational predictions at the molecular level， molecular docking simulations were performed to evaluate the binding affinities and interaction modes between the core active flavonoids and the identified hub targets. The docking results demonstrated that isorhamnetin， luteolin， quercetin， kaempferol， and myricetin all exhibited strong and stable binding conformations within the active catalytic pockets of both SRC and MMP9. These compounds were observed to form critical hydrogen bonds and hydrophobic interactions with key amino acid residues， indicating a favorable thermodynamic profile and a high potential for functional inhibition of these key targets in vivo. The final phase of this investigation involved experimental validation using an established murine model of acute eczema. The model was successfully induced on the dorsal skin of KM mice through repeated topical sensitization and challenge with 2，4-dinitrochlorobenzene. Following model establishment， the therapeutic impact of S. solidaginea administration was rigorously assessed at the macroscopic， microscopic， and molecular levels. Macroscopic observation and scoring of clinical symptoms revealed a significant amelioration of erythema， edema， and excoriation in the treatment groups. Histopathological examination of skin tissue sections stained with hematoxylin and eosin （HE） confirmed that S. solidaginea treatment markedly reduced epidermal hyperplasia， spongiosis， and dermal inflammatory cell infiltration. Mechanistically， quantitative analysis via enzyme-linked immunosorbent assay （ELISA） demonstrated that S. solidaginea treatment effectively down-regulated the serum concentrations of the key pro-inflammatory cytokines tumor necrosis factor-alpha （TNF-α）， interleukin-6 （IL-6）， and interleukin-17 （IL-17）， while concurrently restoring the level of interferon-gamma （IFN-γ）. This modulation of cytokine profiles indicates a beneficial shift toward restoring the T helper type 1 cell/T helper type 2 cell （Th1/Th2） immune balance， which is typically dysregulated in acute eczematous conditions. Most importantly， Western blot analysis of the lesional skin tissue provided direct biochemical evidence that the extract of S. solidaginea significantly inhibited the phosphorylation of phosphatidylinositol 3-kinase （PI3K） and protein kinase B （Akt）， thereby confirming the suppression of the PI3K/Akt signaling pathway in vivo. In conclusion， this methodological approach enables the rapid， effective， and comprehensive characterization of the phytochemical profile of S. solidaginea. Furthermore， this study preliminarily elucidates that the therapeutic mechanism of this medicinal plant in treating acute eczema involves a multi-component and multi-target synergistic action， primarily through the alleviation of the cutaneous inflammatory response and the restoration of immune homeostasis via the inhibition of the PI3K/Akt signaling axis. These findings provide a robust scientific foundation and a valuable reference for the further development， clinical application， and quality control of S. solidaginea.

Monitoring pollutants in human blood is a crucial basis for assessing human exposure levels and health risks. Per- and polyfluoroalkyl substances （PFAS）， organophosphate esters （OPEs）， and their diester metabolites （di-OPEs） are widespread environmental co-contaminants with significant toxic effects， making it crucial to monitor their internal human exposure levels. However， existing studies have predominantly investigated these substances in isolation， lacking comprehensive research that simultaneously quantifies PFAS， OPEs， and di-OPEs in human serum. Based on the pretreatment method of 96-well solid phase extraction columns， this study compared three extraction columns and optimized the pretreatment steps to establish an ultra performance liquid chromatography-high resolution mass spectrometry （UPLC-HRMS） method for 29 PFAS， 17 OPEs， and 5 di-OPEs in human serum samples. Phree PLR 96-Wellplate was used as a cleanup plate. 300 µL of 1% formic acid in acetonitrile， 100 µL of the serum sample， and internal standards were added to the cleanup device in sequence. After standing for 5 minutes， the 96-well plate positive pressure device was used to press the mixture into the 96-well collection plate. Finally， the sample was eluted with 100 µL of 1% formic acid in acetonitrile. The eluate was collected and concentrated for the detection of PFAS， OPEs， and di-OPEs by UPLC-HRMS. PFAS and OPEs were detected using an Acclaim RSLC 120 C18 column， while di-OPEs were detected using an Acquity UPLC BEH C18 column. Both were subjected to gradient elution with methanol and 5 mmol/L ammonium acetate in water as the mobile phases. Sample ionization was performed using a heated electrospray ionization source （H-ESI）. PFAS and di-OPEs were analyzed in negative ion mode， while OPEs were analyzed in positive ion mode. Data acquisition was conducted in full-scan/data-dependent tandem mass spectrometry （Full MS/ddMS²） mode. Quantification was achieved using the internal standard calibration method to ensure measurement accuracy. The results showed that under the optimized conditions， the target compounds had good linear relationships in the range of 0.05–50 ng/mL （R² > 0.99）， and the method detection limits （MDLs） of 29 PFAS， 17 OPEs and 5 di-OPEs were 0.000 120–0.274 ng/mL， 0.011 0–0.250 ng/mL， and 0.012 0–0.220 ng/mL， respectively， and the spiked recoveries were between 45.9% and 147.8%. The relative standard deviations （RSDs） were 1.2%–29.0%. Most PFAS and di-OPEs had matrix enhancement effects， and most OPEs had matrix inhibition effects. Among them， hexafluoropropylene oxide dimer acid （GenX） （196.5%） and trimethylphenyl phosphate （TMPP） （54.6%） had significant matrix enhancement and inhibition effects， respectively， which could be corrected with appropriate internal standards. The recoveries of these two substances after correction were 127.3% and 78.7%， respectively， which met the analysis requirements. The proposed approach offers significant practical benefits， combining straightforward operation with shortened extraction time and enhanced throughput， which was validated through analysis of 15 human serum samples collected in Jinan in 2024. The total contents of 29 PFAS were 6.71–379 ng/mL， with a median value of 22.9 ng/mL. Eight PFAS were detected with a detection frequency of 100.0%， with median contents of perfluorooctanoic acid （PFOA） and perfluorooctane sulfonate （PFOS） reaching 8.96 ng/mL and 4.07 ng/mL， respectively. The total contents of 17 OPEs were 0.015 0–10.5 ng/mL， with a median value of 2.81 ng/mL. The most frequently detected OPEs， with detection frequencies exceeding 60.0%， were triethyl phosphate （TEP）， tri-n-butyl phosphate （TnBP）， and triphenyl phosphate （TPHP）. The total contents of 5 di-OPEs were <MDL–0.443 ng/mL， with a median value of 0.015 0 ng/mL. Therefore， the combined exposure to these pollutants in human blood and its potential health risks demand serious attention.

Pyrethroids are extensively employed in agricultural pest management and household sanitation practices. However， their widespread use has raised concerns as they pose a multitude of health risks to humans. Consequently， the development of precise， highly sensitive， and efficient biomonitoring techniques for evaluating internal exposure levels of pyrethroids across different populations has emerged as a paramount goal in the field of environmental exposure and health effect research. This study developed a method using ultra performance liquid chromatography-tandem mass spectrometry （UPLC-MS/MS） for the simultaneous determination of five common pyrethroid metabolites in urine， with sample pretreatment involving hydrochloric acid hydrolysis and liquid-liquid extraction. The method was optimized for mass spectrometric acquisition parameters and liquid chromatography separation conditions. Chromatographic separation was successfully accomplished utilizing a BEH C₁₈ column （100 mm×2.1 mm，1.7 μm）. Mass spectrometric data were acquired in negative ion mode under multiple reaction monitoring （MRM） conditions. Among the analytes， trans-3-（2，2-dichlorovinyl）-2，2-dimethylcyclopropanecarboxylic acid （trans-DCCA） and cis-3-（2，2-dichlorovinyl）-2，2-dimethylcyclopropanecarboxylic acid （cis-DCCA） are isomers， as are 2-phenoxybenzoic acid （2-PBA） and 3-phenoxybenzoic acid （3-PBA）， and they share identical MS acquisition parameters. Identification was based on reference standards and retention times. The mobile phase consisted of 0.1% acetic acid in water （A） and acetonitrile （B）. The gradient elution program was as follows： 0-0.5 min， 10%B； 0.5-4.5 min， 10%B-70%B； 4.5-5 min， 70%B-100%B； 5-7 min， 100%B； 7-8 min， 100%B-10%B； 8-10 min， 10%B. The optimization of urine sample pretreatment conditions was divided into two parts： hydrolysis and extraction. Using the recoveries of target analytes as the evaluation metric， parameters including the dosage of hydrolysis reagent， hydrolysis temperature and duration， as well as the type， dosage， and extraction time of the extraction solvent were systematically optimized. The optimized pretreatment protocol is delineated as follows： Initially， 40 µL of the 2-PBA internal standard working solution was precisely added to 1 mL of urine sample， followed by thorough mixing to ensure homogeneity. Subsequently， 150 µL of hydrochloric acid （2 mol/L） was introduced to facilitate hydrolysis， which was allowed to proceed at ambient room temperature for a duration of 30 min. Finally， extraction was carried out using 2 mL of ethyl acetate， accompanied by vigorous shaking for 30 min to maximize extraction efficiency. Following centrifugation， the organic phase was separated， evaporated to near dryness， and reconstituted with 1.0 mL of acetonitrile prior to instrumental analysis. The matrix effects were evaluated using pure solvent and matrix-matched standards. Among the five analytes， three exhibited a moderate matrix effect， while two showed a weak matrix effect. The recovery performance using 2-PBA as an internal standard was also assessed. Consequently， a combination of the working curve and the internal standard method was selected for the quantification of the target analytes. Based on this， the methodological parameters of the method were validated. The results indicate that the five pyrethroid metabolites exhibited good linearity， with correlation coefficients of the calibration curves all exceeding 0.995. The limits of detection （LODs） ranged from 0.13 ng/mL to 1.32 ng/mL， and the limits of quantification （LOQs） ranged from 0.44 ng/mL to 4.39 ng/mL. The average recoveries of the samples at three spiked levels of 20， 50， and 80 ng/mL ranged from 91.0% to 102.0%. The intra-batch precision was between 1.1% and 8.1%， while the inter-batch precision was between 1.1% and 4.6%. Sample stability was demonstrated for at least one week when stored at 4 ℃. The established method was applied to analyze 18 urine samples from the general population. Neither cis-DCCA nor 4-fluoro-3-phenoxybenzoic acid （4F-3PBA） was detected in any sample. The mass concentration of 3-PBA ranged from 0.69 ng/mL to 1.59 ng/mL， with a detection rate as high as 88.9%. These results are largely consistent with screening studies on human pyrethroid metabolite levels reported in domestic and international literature in terms of both the detection rate and mass concentration range of 3-PBA. The presence of 3-PBA may originate from household insecticide exposure or dietary sources. The method demonstrates simple and efficient sample pretreatment， strong cost-effectiveness， low LODs， and high accuracy and precision. It can therefore serve as a reliable technical reference for monitoring and exposure assessment in various populations， particularly sensitive groups such as the general population， pregnant women， and children.