1，3-Oleic-2-palmitic triglyceride （OPO） has received widespread attention as a nutritional fortifier allowed to be added to infant formulas， and there is an urgent need to develop high-purity certified reference material （CRM） to meet the metrological traceability requirements of relevant tests. Value assignment methods for pure materials are a key technical challenge in the development of high-purity CRMs. The commonly used mass balance （MB） method indirectly determines the purity of the material by subtracting impurities， which involves the accurate quantification of structurally similar impurities， moisture， volatile solvents， and non-volatile impurities. Among these steps， the separation and analysis of structurally similar impurities of the main component are the key to MB method. Owing to the lack of ultraviolet/fluorescence chromophores， OPO cannot be effectively detected by liquid chromatography with terminal absorption （190–210 nm）； this technique lacks sufficient sensitivity for low-level related impurities. Therefore， with a universal detector， this study established a high performance liquid chromatography-charged aerosol detector （HPLC-CAD） method for the analysis of structural analogue impurities in OPO， and applied the mass balance method to certify the purity of the reference material. On the other hand， based on quantitative nuclear magnetic resonance （qNMR） technology， a qNMR purity assignment method for OPO purity was established by optimizing key parameters such as NMR internal standards and quantitative peaks. In the mass balance method， an HPLC-CAD was selected as the core technique for quantifying trace structurally similar impurities of OPO. The chromatographic conditions were as follows： Nova Pak C8 column （150 mm×3.9 mm， 4 μm）， acetonitrile-isopropanol （90∶10， volume ratio） at a flow rate of 1.0 mL/min， sample dissolved in n-hexane-isopropanol （1∶1， volume ratio） at a mass concentration of 2.0 mg/mL with an injection volume of 5 μL. Under these conditions， four structurally related trace impurities were baseline-separated and accurately quantified. Method validation showed that in the mass concentration range of 0.001-0.025 mg/mL， good linear relationship was obtained （R²=0.999 1）. The limits of detection （LOD） and quantification （LOQ） were 0.000 5 mg/mL and 0.001 mg/mL， respectively， meeting the requirements for quantitative analysis. Six replicate measurements gave an average total impurity content of 1.23% （standard deviation 0.03%）， corresponding to an OPO main component content of 98.77%. In addition， Karl Fischer coulometric titration was used to determine moisture content， headspace gas chromatography-flame ionization detection （HS-GC-FID） was used to analyze residual volatile organic solvents （VOC）， and thermogravimetric analysis was used to determine non-volatile impurities. The purity of OPO was calculated as 98.51% by subtracting related impurities， moisture， VOC and non-volatile residues. In qNMR， direct and accurate quantification of the main component was achieved by optimizing key parameters： deuterated chloroform was selected as the solvent； ethyl paraben was used as the internal standard； selected chemical shifts of 2.33 （OPO） and 6.88 （ethyl paraben） as quantitative peaks. Accurately weigh approximately 10.0 mg of OPO and 3.0 mg of the internal standard in a brown bottle， dissolved in 0.50 mL of deuterated chloroform， and transferred to a 5 mm NMR tube. After six parallel measurements， the average purity of OPO was calculated to be 98.82%， with a standard deviation of 0.21%. Additionally， the homogeneity and stability of the candidate high-purity OPO reference material were evaluated. The uncertainties introduced by value assignment， inhomogeneity， and instability were evaluated. The results showed that the certified purity value of OPO was 98.7%， with a relative expanded uncertainty of 0.7% （k=2，where k is the coverage factor corresponding to a confidence probability of approximately 95% under the normal distribution）. The successful development of this certified reference material provides a traceability standard for relevant detections and fills the gap in OPO purity reference material.

The separation of chiral compounds using common dimension chiral chromatographic columns consumes a large of mobile phase and a long time. This is mainly due to the fact that the inner diameter of common chiral chromatographic column is usually 4.6 mm and the column length is 25 cm. The larger column volume requires more mobile phase to drive the sample through the separation process， and the longer column length increases the migration path of the sample within the column， resulting in an extended separation time. Moreover， the common dimension chiral column is filled more stationary phase which is usually expensive. The micro-packed column， due to their structural characteristics， with inner diameters typically controlled and a few millimeters and significantly shortened column lengths， show broad application prospects in improving separation efficiency， achieving miniaturization of separation instruments， and reducing the consumption of stationary and mobile phases. In this study， amylose tris（3，5-dimethylphenylcarbamate） （ADMPC） was used as the chiral selector. Amylose molecules have a helical structure， and the size of the helical cavity and the chiral environment can be regulated through substituent modification. The introduction of 3，5-dimethylphenylcarbamate groups can enhance the hydrophobic interaction， hydrogen bonding， and π-π stacking between ADMPC and chiral compounds， thereby improving chiral recognition ability. Due to its excellent selectivity and wide applicability， it has become a research hotspot in the field of chiral separation. Two 10-millimeter-long micro-packed columns were prepared by coating ADMPC on silica gel particles with diameters of 500 nm and 1.7 μm， respectively， and were named ADMPC-1 （500 nm silica gel） and ADMPC-2 （1.7 μm silica gel）. On these two micro-packed columns， 23 and 20 racemates， respectively， including various types such as alcohols， esters， ketones， and acids， achieved good separation effects. Notably， there are certain differences in separation selectivity between these two micro-packed columns， showing a certain degree of complementarity. Moreover， these two micro-packed columns also exhibit this complementary property compared with the commercial Chiralpak AD-H chiral column. The most significant advantage of micro-packed columns is the short separation time， most compounds can be separated within 8 min， and many compounds can achieve baseline separation. Commercial chiral columns， due to their longer column lengths， usually require 15-30 min or even longer for sample retention time， and some compounds may not achieve baseline separation due to insufficient resolution. The shorter column length and optimized small size silica gel of micro-packed columns significantly reduce the retention time of samples and improve resolution， which is of great significance for high-throughput screening experiments （such as chiral compound screening in drug development）， and can significantly improve experimental efficiency. At the same time， due to the reduced column volume， the amount of mobile phase used is also very small. Common dimension chiral columns usually consume tens to hundreds of milliliters of mobile phase for one analysis， while the micro-packed columns in this study only require a few milliliters or even less for one analysis. This greatly reduces the generation of organic waste and lowers environmental pollution， in line with the development concept of green chemistry. In addition， compared with common dimension chiral columns， the amount of stationary phase in micro-packed columns is significantly reduced， significantly lowering the cost； at the same time， it can also provide suitable column dimemsion for the development of miniaturized chromatographic systems.

An ion exclusion chromatography method was established for the simultaneous determination of glycerol， acetic acid and sodium bisulfite in compound amino acid injection （14AA）. No tedious sample pretreatment is required； the sample can be directly injected after filtration through a 0.22 μm membrane filter. Chromatographic separation was achieved using an Xtimate Sugar-H column （300 mm×7.8 mm， 5 μm） packed with a strong ion-exchange resin based on a rigid styrene/divinylbenzene matrix， which is specially designed for the separation of sugar alcohols and organic acids via the ion-exclusion mechanism. The mobile phase was 7.5 mmol/L sulfuric acid solution， which was selected as it can suppress the ionization of target analytes and enhance their retention on the stationary phase； the amino groups of amino acid components undergo strong ionization， leading to no retention. Isocratic elution was adopted to ensure stable separation efficiency and short analysis time， with the flow rate set at 0.5 mL/min. Based on the ultraviolet absorption characteristics of the target analytes， 200 nm was selected as the detection wavelength. The column temperature was set at 65 ℃ and the injection volume was optimized to 10 μL， which ensures the detection sensitivity while avoiding column overload. The external standard method was adopted for quantitative analysis， featuring simple operation and reliable results. The method exhibited excellent specificity： blank solvent and negative matrix solution tests verified that amino acids caused no interference with the determination of target analytes， and the target analytes achieved good resolution from adjacent peaks. For the linearity investigation， standard solutions were prepared with linear ranges set as follows： 1.197–59.87 mg/mL for glycerol， 0.036 08–1.804 mg/mL for acetic acid， and 0.000 791 6–0.633 3 mg/mL for sulfite. Within these mass concentration ranges， the linear correlation coefficients （r） of all components were greater than 0.999 5， indicating an excellent linear relationship of the method. The accuracy was verified by spiked recovery tests at three levels （80%， 100% and 120% of the labeled amount）， with three parallel preparations for each level. The results showed that the average recoveries of the target analytes ranged from 99.21% to 102.6%， with relative standard deviations （RSDs） of 0.09% to 0.86% （n=3）， demonstrating the high accuracy of this method. The ion exclusion chromatography method established in this study can effectively eliminate the interference of the amino acid matrix in compound amino acid injection （14AA）， and has the advantages of rapid analysis， simple operation， strong specificity and high accuracy， making it suitable for the routine quality control of glycerol， acetic acid and sodium bisulfite in this preparation. This method provides a reliable analytical and testing tool for the quality assurance and safety control of compound amino acid injection （14AA） during production and storage.

Methyl orange （MO） is a typical water-soluble azo organic dye. It is commonly used as an acid-base indicator in basic experiments and as a common organic dye in industrial production. It has strong polluting effects on water bodies. If azo dyes are accidentally ingested， the azo bond is cleaved by azoreductase， generating highly toxic aromatic amines that pose potential carcinogenic risks. Ionic covalent organic frameworks （ICOFs） are novel materials derived from covalent organic frameworks， featuring charged structures on their frameworks and pores. ICOFs exhibit excellent adsorption performance in aqueous solutions. Their high surface area and abundant porous channels facilitate rapid diffusion of pollutants into the inner structure， enabling effective pollutant adsorption. Eggplant is a sustainable biomass material with regular vertical macropores， which can be used as a substrate. This experiment employed an in-situ solvothermal method to prepare a highly efficient adsorbent monolithic material of eggplant composite aerogel （EP@PDA-TGDha COF）. It is applied to organic dye wastewater treatment. Developing this experiment as an undergraduate teaching module of “I love the laboratory” helps cultivate and enhance students’ abilities in sample pretreatment， material synthesis， structural and property characterization， and practical sample analysis. Students enter the research group offline， experience the academic atmosphere， keep abreast of cutting-edge research， taking the crucial first step in their academic journey. Through the “I Love the Lab” research training program， students significantly enhance their professional knowledge， experimental skills， and research-oriented thinking abilities. It also fosters students’ environmental protection awareness and enables them to apply their knowledge and skills to environmental conservation.

On the basis of previous experiments and research projects， this study developed an undergraduate instrumental analysis experiment by expanding the functionality of existing instruments. The primary objective was to enhance awareness among both instructors and students regarding the importance of sample pretreatment. Using Chrysanthemum as the analytical sample， an analytical method for determining the content of camphor and borneol was established based on multiple headspace extraction-gas chromatography-mass spectrometry （MHE-GC-MS）. By systematically optimizing key parameters such as headspace equilibration temperature and time， the optimal pretreatment conditions were determined as an equilibration temperature of 100 ℃ and an equilibration time of 20 min. Method validation demonstrated that both the liquid and solid external standard methods exhibit good linear relationships， correlation coefficients （R²） ranging from 0.981 2 to 0.999 2. Using the established method， the measured contents of camphor and borneol in Chrysanthemum were 0.917 4 mg/g and 0.944 2 mg/g， respectively. These results are significantly higher than those obtained by traditional hydrodistillation-solvent extraction， reflecting the near-complete extraction advantage of MHE. As a comprehensive inquiry-based teaching project， this experiment has been successfully integrated into the instrumental analysis laboratory course， effectively cultivating students’ comprehensive practical skills and scientific thinking in method development， condition optimization， and data processing. The proposed teaching method does not require additional experimental reagents and demonstrates good universality， extendibility， and promotability， providing valuable insights for the reform of instrumental analysis laboratory teaching.

Mass spectrometry professionals are in short supply industry-wide. Undergraduate teaching has bottlenecks： limited instrumentation， weak qualitative skills and abstract theoretical concepts. We report an innovative pedagogy coupling conventional gas chromatography-mass spectrometry （GC-MS） with the MassWorks^TM software suite. A 14-component fragrance standard mixture was employed to design an experiment spanning the entire workflow separation， detection， exact-mass measurement， isotopic-pattern matching， molecular-formula assignment， structural elucidation. By exploiting the software’s dual-calibration algorithms for exact-mass and isotopic-profile refinement， the mass accuracy and qualitative reliability of GC-low resolution MS data were dramatically enhanced. The practical results show that this method improves the quality accuracy of MS data by 100 times， controls the mass deviation within |Δm|≤10 mDa， achieves an accuracy of over 98% in isotope spectra， and provides accurate and reliable qualitative results. The teaching effect is significant. Through the closed-loop training of theoretical analysis， software application， and practical verification， students’ scientific research thinking， data analysis ability， and innovative ability to solve complex problems have been effectively improved. This provides a replicable and scalable effective paradigm for improving the quality of instrument analysis teaching under limited resource conditions.

Mass spectrometer is an important component of the instrumental analysis courses in higher education. However， conventional commercial mass spectrometers usually lead to an imbalance between theory and practice in teaching， because of their characteristics of large size， high price and complicated operations， which makes it difficult to establish a direct link between instrument structure and detection principles. The miniature mass spectrometer shares the same core principles as commercial mass spectrometers. Equipped with an atmospheric pressure ionization （API） source， it significantly reduces sample preparation requirements， and enables rapid analysis of complex samples. The unique capability of the linear ion trap mass analyzer to scan， store， and eject ions enables a dramatic reduction in instrument size. The miniature ion funnel transmission technology and atmospheric pressure continuous sampling interface ensure high repeatability and rapid analysis， allowing compatibility with various atmospheric pressure ion sources for diverse sample types. In terms of performance， the miniature mass spectrometer has a mass-to-charge （m/z） scan range of 50-2 000， and supports tandem mass spectrometry （MSⁿ）， enabling structural analysis of common organic compounds and rapid testing of complex samples. Therefore， the miniature mass spectrometer becomes an ideal tool for experimental teaching. This teaching course has developed a “three-dimensional integrated” teaching system aimed at enhancing students’ knowledge， abilities， and literacy. Firstly， students disassemble key components of the instrument such as the ion source， ion funnel and trap to understand how they work together to achieve sample ionization， transmission， and acquisition. Mass calibration and full scan test help students master the basic operation. By adjusting the radio frequency （RF） voltage parameters， students could observe the impact of different parameters on the experimental results. Secondly， selected ion monitoring and tandem MS analysis help enhance students’ data analysis skills and further develop their abilities in structural confirmation of compounds with MS. Lastly， pharmaceutical ingredient analysis， blood drug testing and multi-component screening of traditional Chinese medicine comprehensively demonstrate the wide applications of MS in scientific research and daily life， aiming to cultivate students’ research thinking and problem-solving abilities. The gradient experiments （from basic calibration to advanced MSⁿ） combined with practical applications have effectively driven the implementation of the offline teaching course. The suggested experimental class hours are 12， covering pre-class preparation， in-class hands-on practice， data acquisition， and post-class summary. Practice has demonstrated that this integrated “structure-principle-application” teaching model effectively bridges the knowledge chain from theoretical learning to practical application， and significantly enhances students’ innovative thinking and ability to solve complex analytical problems， better preparing them to meet the requirements of modern scientific research and industrial applications. Additionally， instructional reflections and feasible teaching optimizations are necessary based on the teaching effects and feedback. This comprehensive learning experience provides students with invaluable inspiration and guidance， and lays a solid foundation for them to enter the field of scientific research， having successfully cultivated talent with practical abilities and innovation literacy， and achieving the intended teaching objectives.

High performance liquid chromatography （HPLC） remains a core separation and analytical technique in modern analytical chemistry. Boasting prominent advantages including high quantitative accuracy， rapid analysis speed， strong selectivity， and high sensitivity， it has deeply permeated key fields such as biochemistry， pharmaceutical R&D， food testing， environmental monitoring， and materials science， delivering crucial support for the accurate qualitative and quantitative analysis of target components in complex systems. However， with the increasing complexity of analytical samples in scientific research and industrial production， marked by enhanced matrix interference， an expanded polarity range of target compounds， and widespread coexistence of multiple components， the limitations of traditional single-mode chromatography （e.g.， reversed-phase liquid chromatography （RPLC）， hydrophilic interaction chromatography （HILIC）， ion-exchange chromatography （IEC）） have become increasingly pronounced. Specifically， RPLC exhibits insufficient retention and separation capabilities for highly polar compounds， HILIC struggles to handle hydrophobic substances effectively， and IEC is only applicable to the separation of ionic components. None of these single-mode techniques can meet the demand for efficient and comprehensive separation of complex samples. To address this technical bottleneck， mixed-mode chromatography （MMC） has emerged as a viable solution. Its core innovation lies in integrating two or more separation mechanisms into a single chromatographic column. Through the synergistic effects of functionalized stationary phases， MMC enables efficient separation and accurate analysis of complex systems， thereby significantly expanding the application scope of HPLC. This paper briefly elaborates on the separation mechanisms of four mainstream mixed modes， namely RPLC/IEC， RPLC/HILIC， HILIC/IEC， and RPLC/HILIC/IEC. It also summarizes in detail the key chemical reaction types for stationary phase preparation （e.g.， click chemistry reactions and free radical polymerization reactions）， typical packing structures， as well as the characteristics and preparation strategies of novel functional materials such as porous organic cages （POCs）， metal-organic frameworks （MOFs）， covalent organic frameworks （COFs）， carbon quantum dots （CQDs）， microporous organic networks （MONs）， and ionic liquids （ILs）. Based on the latest research findings from 2020 to 2024， this paper systematically reviews the application cases of the aforementioned four mixed-mode stationary phases in practical scenarios such as traditional Chinese medicine component analysis， environmental pollutant detection， food quality control， and pharmaceutical research and development. It also conducts an in-depth analysis of the technical advantages of these four mixed modes， as well as the limitations of some stationary phases， including insufficient stability under extreme pH conditions， complex preparation processes， and high costs for large-scale production. Finally， this paper outlines the core challenges currently confronting mixed-mode stationary phases， including cumbersome synthesis steps， easy degradation and inactivation of functional groups， and difficulties in mobile phase optimization. It further points out that future development trends should focus on simplifying preparation processes， developing environmentally friendly and smart responsive materials， and enhancing the feasibility of large-scale production. The aim is to provide theoretical reference and technical support for the design and development of novel high-efficiency stationary phases， and facilitate greater breakthroughs in mixed-mode chromatography technology in the field of complex sample separation.

Carrimycin is a macrolide antibiotic widely used in the pharmaceutical and clinical fields， with its primary composition consisting of three distinct derivatives of spiramycin. For decades， macrolides like carrimycin have been valued for their efficacy against Gram-positive bacteria and certain atypical pathogens. However， as research has progressed， recent studies have revealed that carrimycin， along with various other structurally modified spiramycin derivatives， exhibits significant antitumor activity in multiple experimental models. This finding suggests that the spiramycin molecular scaffold possesses intrinsic potential beyond its conventional antibacterial role. By strategically modifying the substituents attached to this core structure， it may be feasible to develop novel compounds with enhanced and more selective antitumor properties. Despite this promising outlook， research into the specific targets and comprehensive mechanisms underlying the antitumor effects of spiramycin derivatives remains notably insufficient. A major gap exists in the precise identification of their molecular targets within cancer cells and the detailed signaling pathways they modulate. This lack of mechanistic understanding poses a substantial barrier to the rational design， optimization， and clinical translation of spiramycin-based antitumor therapeutics. To systematically address this knowledge gap and elucidate the antitumor mechanism of this compound class， the present study adopted an activity-based protein profiling （ABPP） strategy. ABPP is a chemoproteomic approach that enables the direct identification of functionally active proteins that interact with small-molecule probes in a native biological context. As a first step， we designed and synthesized a novel， high-activity spiramycin derivative termed n-hexanoyl spiramycin （h-SPM）. Building upon the structure of h-SPM， we subsequently engineered and synthesized a structurally analogous activity-based probe. This probe was specifically functionalized with chemical handles （such as an alkyne group） compatible with ABPP methodologies， allowing for downstream bioorthogonal conjugation and enrichment steps. The experimental workflow began by incubating this active probe with live cancer cells. During this co-incubation period， the probe engaged with and covalently bound to its potential protein targets within the complex cellular environment. Following the interaction， cells were lysed， and the probe-labeled proteins were efficiently isolated and purified using affinity-based enrichment techniques—specifically via copper-catalyzed azide-alkyne cycloaddition （click chemistry） to immobilize them onto a solid support. The enriched protein pool was then subjected to in-depth analysis using liquid chromatography-mass spectrometry （LC-MS）. This analytical phase yielded detailed， proteome-wide information on the identities of proteins that interact with the h-SPM-based probe. To extract biological insights from the list of identified proteins， we performed comprehensive bioinformatic analysis using Gene Ontology （GO） enrichment. This systematic classification provided crucial information regarding the biological processes， molecular functions， and cellular components associated with the captured proteins. The functional annotations derived from GO analysis allowed us to evaluate and prioritize several promising candidate target proteins for further experimental validation. Through this integrated proteomic and bioinformatic approach， we successfully identified multiple potential cellular targets of h-SPM. Notable among these were amyloid precursor protein （APP） and low-density lipoprotein receptor （LDLR）， both of which are implicated in diverse cellular processes such as cell adhesion， signal transduction， and metabolic regulation. To confirm the biological relevance of these interactions， we employed Western Blotting experiments. These studies verified that the expression or post-translational modification states of APP and LDLR were altered in response to h-SPM treatment， thereby confirming their status as responsive molecular targets. After identifying and preliminarily validating APP as a key interactor， we proceeded to investigate its functional role in the drug’s mechanism of action. Using short hairpin RNA （shRNA）-mediated protein knockdown， we generated cancer cell lines with significantly reduced APP expression. Comparative analysis of drug sensitivity between these knockdown cells and their wild-type counterparts revealed a marked attenuation of h-SPM’s antitumor effects in the absence of APP. Complementary to this， cell staining assays， including immunofluorescence， were conducted to visualize morphological changes， alterations in cell viability， and the subcellular localization of relevant biomarkers following h-SPM treatment. Collectively， these functional experiments provided compelling evidence that APP plays a critical and indispensable role in mediating the antitumor activity of h-SPM. Our findings thus outline a preliminary model of the drug’s mechanism， likely involving cellular pathways regulated or influenced by APP. In summary， this study achieves two significant objectives. First， it establishes and validates a robust， generalizable target-screening platform based on ABPP， specifically tailored for the investigation of spiramycin-class compounds. Second， and more importantly， it delivers novel biological insights by pinpointing specific protein targets such as APP. The identification of these targets provides invaluable mechanistic clues and a solid theoretical foundation for the future development of this family of compounds. This work effectively advances spiramycin derivatives from compounds with observed phenotypic activity toward agents with an emerging mechanistic understanding， thereby paving the way for more targeted drug design and informed combination therapy strategies in oncology. Future studies will focus on delineating the detailed downstream consequences of APP engagement by h-SPM and exploring the therapeutic potential of other identified targets in preclinical models.

Quality control of traditional Chinese medicine （TCM） has always been a key and challenging issue in the field of its modernization research. This has posed a high demand for advanced separation materials due to the complex compositions. Covalent organic framework materials （COFs） are a new class of porous crystalline materials composed of multidentate organic units connected by covalent bonds. They have demonstrated significant application value in areas such as catalysis and chromatographic analysis. This study focused on developing a novel core-shell-type chromatographic stationary phase using 2，4，6-tris（4-aminophenyl）-1，3，5-triazine （TAPT） and 1，4-benzenedicarboxaldehyde （TA） as building units. The TAPT-TA-COF@SiO₂ core-shell composite materials were successfully prepared on the surface of silica microspheres using a multi-step polymerization strategy， in which the SiO₂ cores were fabricated using the polymerization-induced colloidal aggregation （PICA） method. The imine-linked TAPT-TA-COF@SiO₂ core-shell stationary phase was subjected to comprehensive physicochemical characterization and chromatographic evaluation experiments. The analytical techniques employed included scanning electron microscopy （SEM）， transmission electron microscopy （TEM）， energy dispersive spectroscopy analysis （EDS）， nitrogen adsorption-desorption isotherms， Fourier transform infrared spectroscopy （FT-IR）， and powder X-ray diffraction （PXRD）. The systematic characterization results clearly indicate that the prepared stationary phase exhibits excellent monodispersity， and the COF layer is uniformly coated on the surface of the SiO₂ core. TEM characterization demonstrated that the thickness of the COF materials on the surface of SiO₂ is approximately 110 nm. Furthermore， FT-IR spectra were collected and the results demonstrated that the characteristic stretching vibrations at 3 209， 2 927， and 1 515 cm^-1， attributed to N-H， C-H， and C=N stretching， confirm the condensation reaction between TAPT and TA. In the XRD pattern， the peaks observed at 16.6°， 18.9°， 25.2° and 27.5° were attributed to the COF material and were consistent with previous reports， thereby confirming the successful synthesis of this derivative. The N₂ adsorption-desorption isotherm analysis confirmed that the material possesses a typical mesoporous structure. Its specific surface area and pore size distribution are similar to those of the original porous SiO₂ microspheres， subsequently providing a structural basis for efficient chromatographic mass transfer kinetics. In addition， the chromatographic performance was investigated. And it was confirmed that the stationary phase was effectively used for the separation of representative neutral polar or non-polar compounds such as benzenes， alkylbenzenes， phthalate esters， formamides， and aniline mixtures. These compounds are separated due to hydrophobic interactions， π-π interactions， and the unique mesoporous structure in the reversed-phase chromatography mode. ACN-water （30∶80 or 40∶60， volume ratio） was selected as the mobile phase at a flow rate of 1 mL/min. The results of the methodological validation indicate that the intra-batch relative standard deviations （RSDs） of one TAPT-TA-COF@SiO₂ packed chromatographic column were less than 1.6%， demonstrating excellent preparation reproducibility. Furthermore， this stationary phase was applied to the quality control of traditional Chinese medicine. Specifically， it was used to determine the content of astragaloside Ⅳ in Astragalus according to pharmacopoeial records. The measured result met the pharmacopoeia standard of ≥0.08%. This research work not only provides new opportunities for advancing fundamental and applied research on novel COF stationary phases， but also helps to further promote in-depth research on the application of COF materials at the intersection of separation science and pharmaceutical sciences.