分子印迹技术是在模板分子(印迹分子)存在下人工制备分子印迹材料的技术,由于该技术在分子识别领域具有结构预定性、高效识别性和广泛实用性的特点,自现代分子印迹奠基人 Guenter Wulff和 Klaus Mos-bach 开创性地提出"共价印迹法"(1972 年)[1]和"非共价印迹法"(1993年)[2]以来,分子印迹技术已经在色谱分离、固相萃取、仿生传感、模拟酶催化、临床药物分析等领域取得了长足的进步.分子印迹材料的核心是识别位点,在共价印迹中,可以选用的可逆共价键有限;在非共价印迹中,常用的非共价键作用力(范德华力、氢键作用力等)存在结合力不足的问题.因此,协同分子印迹技术应运而生,即在印迹位点内,通过多种作用力(如后修饰小分子配基、核酸适配体等)实现对目标分子的高效识别.
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 MassWorksTM 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.
In analytical chemistry， separation and enrichment steps serve not only as critical stages in the analytical workflow， but also as core technologies that determine the accuracy， reliability， and applicability of analytical methods. However， in conventional experimental instruction， this step is frequently overlooked owing to time constraints and an overemphasis on instrumental techniques， leading to fragmented knowledge structures and hindering the development of integrated analytical thinking that connects theory with practice. To address this issue， this study， based on the "knowledge-ability-practice" three-dimensional integrated teaching objective， innovatively establishes a "positive-negative case-based teaching" model， using the solid-phase microextraction （SPME） of six nitrogen-containing pesticides from environmental water samples as the instructional case. By comparing fibers coated with non-polar polydimethylsiloxane （PDMS） and strongly polar polyacrylate （PA）， and employing high performance liquid chromatography with ultraviolet detection （HPLC-UV）， this study systematically investigates the matching relationship between analyte polarity and coating properties. In the positive case， the PDMS coating demonstrates superior extraction performance for weakly polar pesticides based on the "like dissolves like" principle， with a linear coefficient of determination R2≥0.993 7 and detection limits ranging from 0.019 to 0.17 μg/L. In contrast， in the negative case， the PA coating shows weaker extraction efficiency due to differences in the extraction mechanism， yielding detection limits between 0.066 to 1.069 μg/L. Based on spiked recovery tests conducted on actual river water samples from Lanzhou （recoveries： 81.5% to 117%）， a multi-dimensional evaluation system of "knowledge internalization， competency enhancement， and practical literacy" is established accordingly. Teaching feedback demonstrates that the students show a marked improvement in their understanding of the core principles， and their innovative thinking and problem-solving skills are effectively cultivated. The "contrasting-case-based" pedagogical approach effectively bridges theory and experiment while stimulating students' investigative thinking. The study provides a replicable and scalable practical model for the reform of analytical chemistry experimental teaching， which holds significant value in cultivating high-caliber talents with rational method selection and problem-solving abilities.
This paper designs an experimental scheme to meet the undergraduate experimental teaching requirements of the Pharmaceutical Engineering major. The content focuses on the detection of bifenthrin pesticide residue in tea by QuEChERS-GC-MS. The experimental contents include sample pretreatment， single-factor screening and response surface methodology optimization of QuEChERS experimental conditions， establishment of a GC-MS analysis method， determination of bifenthrin pesticide residues in tea samples， and experimental data processing. It aims to cultivate students' ability to comprehensively apply modern analytical techniques to solve practical problems. Through this experiment， students can master the basic principles and operational skills of GC-MS instruments and equipment， and understand the complete process of pesticide residue analysis in real samples， and recognize the importance of sample pretreatment for the analysis of trace components in complex systems. Under the QuEChERS experimental conditions screened in the single-factor experiment， the recovery rate of bifenthrin in the tea samples was 85.9%. Under the QuEChERS experimental conditions optimized by the response surface methodology， the recovery rate of bifenthrin in tea samples can reach 90.33%. Among the 38 tea samples， bifenthrin was detected in 12 samples， with the maximum detected value of 0.911 mg/kg. None of them exceeded the maximum residue limit of bifenthrin in tea stipulated in the National Food Safety Standard GB 2763-2021. This teaching design involves learning relevant content before the experiment， training operational skills during the experiment， and conducting data analysis after the experiment. It can not only stimulate students' interest in learning， but also cultivate their rigorous scientific thinking. It enables students to proactively face problems， actively analyze them and try to solve them throughout the entire learning process.
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.
Current undergraduate environmental chemistry curricula often suffer from a disconnect between sample preparation and large-scale instrumental analysis， making it difficult to cultivate students' systematic analytical capabilities. Furthermore， the experimental content predominantly focuses on conventional water quality parameters and traditional pollutants like heavy metals， lacking training in monitoring and remediation technologies for new pollutants， which creates a significant gap with national strategic needs. To address this issue， this study designed an integrated comprehensive experiment that combines material synthesis， sample pretreatment， and large-scale instrumental analysis， targeting typical new pollutants， such as organic ultraviolet filters （OUVFs） from pharmaceuticals and personal care products. A magnetic covalent organic framework （MCOF） was synthesized via a reflux heating method. Students were guided to apply it as the sorbent in magnetic solid-phase extraction （MSPE）. Owing to its facile magnetic separation， the rapid enrichment of OUVFs was achieved within 8 min， followed by quantitative determination of seawater samples using ultra-high performance liquid chromatography-tandem mass spectrometry （UHPLC-MS/MS）. Through a three-stage teaching module encompassing "pre-class preview， in-class experiment， and post-class discussion"， the experiment emphasizes enhancing students' independent exploration abilities in adsorbent synthesis and characterization， optimization of enrichment parameters， and data analysis. Teaching practice has shown that this design not only enabled students to deeply understand the complex sample pretreatment process and the mechanism of multi-factor synergistic effects， but also significantly enhanced their comprehensive innovation and practical abilities in solving new pollutant monitoring problems.
Organic phosphate esters （OPEs） and phthalate esters （PAEs） are two classes of synthetic compounds widely incorporated into various industrial and consumer products as flame retardants and plasticizers. Both exhibit multi-organ cumulative toxicity. In this study， an ultra performance liquid chromatography-tandem mass spectrometry （UPLC-MS/MS） method was developed for the determination of 22 OPEs and their diester metabolites （mOPEs） and 11 phthalate ester metabolites （mPAEs） in whole blood. The whole blood samples were extracted with 0.1% formic acid-acetonitrile by liquid-liquid extraction for protein precipitation， and then purified by HMR S-micro BIO 96-well solid phase extraction column. The filtrate was blown to near dryness with nitrogen， and reconstituted with 0.1% formic acid-acetonitrile solution. The target compounds were detected by UPLC-MS/MS. A Ghost Trap DS-HP chromatographic column （30 mm×2.1 mm） was used as the background ghost peak trap column， and an ACQUITY UPLC® BEH C18 chromatographic column （150 mm×2.1 mm， 1.7 μm） was used as the analytical column. Methanol and 0.5 mmol/L ammonium acetate were used as mobile phase for gradient elution. Electrospray ionization （ESI） was used for MS detection， with both positive and negative ion scanning in the multiple reaction monitoring （MRM） mode. Isotope internal standard method was used for quantification. The results showed that 33 target compounds could be separated within 15 min， and good linear relationships were observed for all target compounds across their respective mass concentration ranges （r=0.993 3-0.999 8）. The limits of detection were 0.003-0.31 ng/mL and the limits of quantification were 0.01-1.02 ng/mL. The spiked recoveries of 33 targets in whole blood samples ranged from 60.5% to 138.3%， with relative standard deviations （RSDs） from 1.6% to 11.0%. The detection rates of tris-（2-chloroisopropyl）phosphate （TCiPP）， triphenyl phosphate （TPHP）， 2-ethylhexyl diphenyl （EHDPP）， mono（2-ethyl-5-hydroxyhexyl） phthalate （MEHHP）， mono-n-butyl phthalate （MNBP） and mono-n-octyl phthalate （MOP） were 100% in 14 whole blood samples， and the highest detection amount of TPHP was 10.04 ng/mL. The developed method is simple to operate， sensitive， accurate， requires small sample volumes， and is cost-effective on reagents. It is suitable for human biomonitoring， and provides a theoretical basis for monitoring organic phosphate and phthalic ester substances in the population.
p-Phenylenediamine compounds （PPDs） are widely used as anti-aging agents and antioxidants in rubber industry. However， environmental concerns have arisen regarding the toxicity of their quinone derivatives （PPD-Qs）. Fine particulate matter （PM2.5） serves as a crucial vector enabling PPDs and PPD-Qs to enter the human body via respiratory exposure pathway. Therefore， accurate monitoring of PPDs and PPD-Qs in PM2.5 is essential. In this research， a high performance liquid chromatography-tandem mass spectrometry （HPLC-MS/MS） approach was established to quantify seven PPDs and seven PPD-Qs in PM2.5. Samples were collected on quartz fiber filters with a diameter of 90 mm. The filter was cut into strips and subsequently spiked with 2 ng of 6PPD-d5 and 6PPD-Q-d5 internal standards. The target compounds were extracted with 10 mL of acetonitrile containing 1% （volume fraction） ammonia solution and 20 μmol/L glutathione （GSH）， and purified via a modified QuEChERS pretreatment method. Specifically， after salting out with 0.5 g of NaCl and 1.0 g of MgSO4， the supernatant was subjected to dispersive solid-phase extraction for cleanup. In this process， 0.5 g of MgSO4， 100 mg of octadecylsilane （C18）， and 100 mg of N-propylethylenediamine （PSA） were employed as sorbents. Subsequently， 5 mL of the cleaned-up supernatant was evaporated to near dryness and then redissolved in 0.2 mL of a methanol-water mixture （1∶1， volume ratio） for instrumental determination. The compounds were separated on a Waters ACQUITY UPLC HSS T3 column（100 mm×2.1 mm，1.8 μm） with mobile phases consisting of a 5 mmol/L ammonium formate aqueous solution containing 0.1% formic acid and methanol. Identification and quantification of the compounds were carried out under positive electrospray ionization in multiple reaction monitoring mode. The results indicated that all analytes exhibited good linear relationships within the range of 0.05-20.0 ng/mL， with correlation coefficients （r） exceeding 0.999. The method detection limits （MDLs） and method quantification limits （MQLs） of the compounds were within the ranges of 0.003-0.07 pg/m3 and 0.01-0.2 pg/m3， respectively. At three spiked levels of low， medium and high， the recoveries of the compounds were in the range of 66.3% to 119.0% ， and the relative standard deviations （RSDs） ranged from 1.5% to 13.2% （n=6）. Ultimately， the developed method was employed for the analysis of PPDs and PPD-Qs in PM2.5 samples collected from Baotou during January and February 2025. The results indicated that seven PPDs and five PPD-Qs were detected， with mass concentrations ranging from 2.26 to 251.2 pg/m3 and 2.36 to 105.4 pg/m3， respectively. The proposed method is characterized by its simplicity， environmental friendliness and accuracy， making it suitable for the rapid quantitative analysis of PPDs and PPD-Qs in PM2.5.
Carbonyl compounds （CCs） are a pivotal class of oxygenated volatile organic compounds in ambient air， which are mainly derived from natural emissions， primary emissions from anthropogenic sources， and secondary formation due to atmospheric oxidation. Various classes of CCs have different impacts on human health， and play critical roles in atmospheric photochemical reactions， ozone production， and the formation of secondary organic aerosols. At present， the number of CCs in ambient air determined by traditional methods is limited， especially for high molecular weight monocarbonyl compounds （HMW-MCs） and dicarbonyl compounds （DCs）. If more CCs can be accurately determined， it will be beneficial for the study of atmospheric oxidation mechanisms and the formulation of pollution control strategies.In this study， an analytical method was developed that enables simultaneous determination of the 40 CCs in ambient air with a single injection， utilizing high-performance liquid chromatography-triple quadrupole mass spectrometry. First， the 16 CCs-2，4-dinitrophenylhydrazine （DNPH）-derivatized single standards were prepared using purified DNPH crystals in the laboratory. Second， the remaining 24 commercial CCs-DNPH-derivatized hybrid standards were purchased. Finally， all the above standards were mixed in a certain proportion to prepare the 40 CCs-DNPH-derivatized hybrid standards for later use. The sample collection procedures （sampling flow rate and sampling duration）， the pretreatment methods （elution volume and constant volume）， the chromatographic analysis conditions （chromatographic column， the concentration of ammonium acetate， etc.） and the mass spectrometry analysis conditions （ion source parameters， fragmentation voltage， collision energy， etc.） were optimized. Ambient air samples were collected using DNPH cartridges （at a flow rate of 1 L/min， for 4 h）， eluted with 4 mL of acetonitrile and diluted to a final volume of 5 mL （the acetonitrile naturally flowed through the DNPH cartridges in the opposite direction to the sampling direction） in a vacuum drying chamber， and then filtered through a 0.22 μm organic filter membrane and transferred to 1.5 mL brown sample bottles prior to instrumental analysis. Chromatographic separation was performed using a Sepax BR-C18 column （250 mm×4.6 mm， 5 μm） with mobile phases consisting of 1.0 mmol/L ammonium aqueous solution and acetonitrile under gradient elution conditions. The analysis was performed at a column temperature of 50 ℃ with an injection volume of 5 μL. The target analytes were identified using negative electrospray ionization （ESI-） and multiple reaction monitoring （MRM） modes and quantified using the external standard method. The results showed that the 40 CCs exhibited good linearities within the mass concentration range of 2.5-200 μg/L， with the determination coefficients （R2） exceeding 0.997 8. The method detection limits and method quantification limits were established within the ranges of 0.001-0.03 μg/m3 and 0.005-0.12 μg/m3， respectively. The blank spiked recoveries of the target analytes at low， middle， and high concentration levels （0.05， 0.1， and 0.2 μg） ranged from 75.2% to 119.0%， while the relative standard deviations （RSDs， n=7） ranged from 0.4% to 4.2%. This analytical method was applied to the monitoring of ambient samples in the Pearl River Delta region. Except for 2-furaldehyde and isophorone， 38 CCs were effectively detected. Among these， formaldehyde， acetaldehyde and nonanal consistently showed higher concentrations across all sampling sites. Overall， the concentrations of low molecular weight monocarbonyl compounds （LMW-MCs） were moderately higher than those of HMW-MCs， and substantially higher than those of DCs. Compared with conventional methods， this method effectively overcame the limitations in existing approaches regarding the limited variety and detection difficulties of DCs and HMW-MCs. Additionally， it also improved the chromatographic resolution of target analytes which have similar retention times and the same ion pairs. Overall， this method was simple and rapid， offering excellent sensitivity， precision， and chromatographic resolution. It was suitable for the determination of the abovementioned 40 CCs in ambient air．.
A supercritical fluid chromatography （SFC） method coupled with UV detection was developed for the separation of linagliptin and its S-enantiomer. The method was validated and successfully applied to detect the S-enantiomer in real pharmaceutical samples. The separation of the enantiomer was investigated using six different chromatographic columns， and different cosolvents were studied. Chromatographic conditions， including column temperature， backpressure， and flow rate， were optimized. The DAICEL CHIRALPAK AD-H column （250 mm×4.6 mm， 5 μm） was used for separation. Supercritical CO2 served as mobile phase A， and ethanol-isopropanol （1∶1， volume ratio） containing 0.25% diethanolamine and 0.25% trifluoroacetic acid was used as mobile phase B. Isocratic elution was carried out at a ratio of A∶B=73∶27 （volume ratio） with a flow rate of 1.5 mL/min. The column temperature was set at 40 ℃， back pressure at 15 MPa， injection volume at 6 μL， and detection wavelength at 220 nm. Under these conditions， linagliptin and its S-enantiomer were separated with a resolution of 3.1 and good peak shapes. Both linagliptin and its S-enantiomer exhibited good linearity in the concentration range of 2-90 μg/mL， with correlation coefficients of 0.999 7 and 0.999 9 （n=8）， respectively. The limit of detection （LOD） for both was 0.8 μg/mL （S/N=3）， and the limit of quantification （LOQ） for both was 2 μg/mL （S/N=10）. The average recoveries of the S-enantiomer spiked at low， medium， and high concentrations in active pharmaceutical ingredients and tablets were 97.4% （RSD=1.1%， n=9） and 101.6% （RSD=1.2%， n=9）， respectively. S-Enantiomer was not detected in three batches of active pharmaceutical ingredients or in three batches of tablets from two different manufacturers. This study represents the first application of SFC for the separation of linagliptin and its S-enantiomer. The method is environmentally friendly， sensitive， and highly efficient， offering good repeatability of peak area. It provides a solid foundation for the quality control of linagliptin and the inclusion of the SFC method in pharmaceutical quality standards， while also offering a useful approach for the rapid separation and impurity control of other chiral drugs.
Alkaloids represent a class of naturally-occurring nitrogen-containing compounds widely distributed across diverse plant species. Owing to their well-documented potential to induce adverse effects on human health， certain alkaloids are explicitly prohibited from being used in cosmetic formulations. The escalating global popularity of essential oil-based cosmetics， which commonly incorporate complex botanical extracts， presents a potential avenue for the inadvertent introduction of these prohibited substances. Consequently， the development of robust， sensitive， and efficient analytical methods for their monitoring is of utmost significance for consumer safety and regulatory compliance. This study devises a reliable， high-throughput approach for the simultaneous determination of 13 prohibited alkaloids in essential oil-based cosmetics. It integrates optimized QuEChERS sample preparation with ultra performance liquid chromatography-tandem mass spectrometry （UPLC-MS/MS）. The sample preparation procedure was meticulously designed to maximize efficiency and minimize analyte loss. Chromatographic separation was accomplished on a Waters ACQUITY UPLC HSS T3 column （100 mm × 2.1 mm， 1.8 μm） maintained at a constant temperature of 30 ℃. The mobile phase was composed of （A） acetonitrile and （B） 0.1% （volume fraction） formic acid aqueous solution. A gradient elution program was implemented at a stable flow rate of 0.3 mL/min according to the following profile： an initial 5， a linear increase to 15 （0-2 min）， a rapid rise to 70 （2-4.5 min）， followed by an immediate reversion to the initial 5 （4.5-5.5 min）， which was maintained for re-equilibration until 7.0 min. The injection volume was 5 µL. Detection and quantification were conducted using a triple quadrupole mass spectrometer equipped with an electrospray ionization （ESI） source operating in positive ion mode （ESI+）. Data acquisition was carried out in the multiple reaction monitoring （MRM） mode to ensure superior specificity and sensitivity. For each of the 13 alkaloids， two specific ion transitions were monitored： one for quantitative analysis and the other for confirmatory identification. The method was rigorously validated in accordance with accepted analytical guidelines. All 13 target alkaloids displayed excellent linearity within a mass concentration range of 0.2 to 50 ng/mL， with correlation coefficients （R2） consistently exceeding 0.99. The limits of detection （LODs） and limits of quantification （LOQs）， determined with acceptable accuracy and precision， ranged from 1 µg/kg to 4 µg/kg and 2 µg/kg to 10 µg/kg， respectively. Method accuracy and precision were assessed through recovery tests at three spiking levels， with six replicates at each level. The mean recoveries for all analytes ranged from 83.9% to 119.1%， with associated relative standard deviations （RSDs） all being ≤7.3%， validating the method's high reliability and repeatability. Systematic evaluation indicated that the matrix effects for the 13 analytes were negligible； hence， the solvent calibration curve was adopted for quantitative analysis. The practical applicability of the validated method was demonstrated through the analysis of 50 batches of commercially available essential oil-based cosmetics. This market survey encompassed 15 products specifically marketed for infants or children and 35 products intended for adult use. As a result， none of the 13 target prohibited alkaloids were detected in any of the tested samples above their respective LOQs. A particularly notable accomplishment of this work is the successful development of a sensitive and reliable quantification strategy for oleandrin， a potent cardiotoxic alkaloid for which standardized detection methods in complex cosmetic matrices such as essential oils have been conspicuously absent. In conclusion， this study successfully establishes a simple， rapid， sensitive， and robust QuEChERS-UPLC-MS/MS method. It is comprehensively validated and clearly well-suited for the routine screening， risk monitoring， and quality control of 13 prohibited alkaloids in a wide array of essential oil-based cosmetics. The method offers reliable technical support for regulatory bodies to enforce safety standards and for manufacturers to ensure the safety and compliance of their products， thereby effectively contributing to the protection of consumer health.
Organic ultraviolet absorbers （OUVAs） are a class of emerging contaminants that have garnered significant attention in recent years. In response to the characteristics of high detection frequency and low concentrations of OUVAs in water bodies， this study established an analytical method based on solid phase extraction （SPE） coupled with ultra-high performance liquid chromatography-tandem mass spectrometry （UHPLC-MS/MS） for the simultaneous determination of 16 OUVAs （nine UV stabilizers and seven UV filters） in drinking water. The 500 mL water sample was pretreated with 25 mg ascorbic acid， acidified to pH 2-3， and spiked with 1 mL of 5 μg/L mixed internal standard working solution. Target compounds were enriched and concentrated using an HLB solid phase extraction cartridge （200 mg/6 mL）. Separation was achieved on an Acquity Premier BEH C18 column （100 mm×2.1 mm， 1.7 μm） using a gradient elution with methanol and 2 mmol/L ammonium acetate aqueous solution as the mobile phases. Detection was performed using electrospray ionization in positive mode and multiple reaction monitoring （MRM）， with quantification carried out by the internal standard method. The precision and accuracy of the established method were evaluated using drinking water as the matrix. The results showed that all 16 OUVAs exhibited good linearity within their respective mass concentration ranges， with correlation coefficients （r） greater than 0.992. The method detection limits （MDL， S/N=3） were 0.03-5 ng/L， and the method quantification limits （MQL， S/N=10） were 0.1-15 ng/L. At spiked levels of 5， 20 and 50 ng/L， the recoveries of the target analytes ranged from 75.0% to 130.6%， with relative standard deviations （RSDs） ranging from 0.9% to 12.9% （n=6）. A total of seven source water samples and seven drinking water samples were analyzed using this method. The results showed that two OUVAs， 2-hydroxy-4-methoxybenzophenone （UV-9） and octocrylene （OC） were detected. In source water， the mass concentrations of UV-9 ranged from <MQL to 13.4 ng/L， while OC mass concentrations ranged from 5.0 to 32.23 ng/L. In drinking water， UV-9 was detected at 0.4 ng/L， and OC mass concentrations ranged from <MQL to 13.2 ng/L. This method is suitable for the trace analysis of 16 OUVAs in drinking water， demonstrating good accuracy and precision. It effectively improves the detection efficiency of OUVAs in water and can be used for the environmental monitoring and analysis of OUVAs in drinking water in China.
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.
Toxic alkaloids,especially ergot alkaloid toxins,present in forage grass and pose serious health hazards to humans and livestock.Hence,a method for simultaneously detecting these dangerous plant toxins is needed.This study established a rapid method for detecting ten ergot alkaloid toxins using solid-phase supported liquid-liquid extraction technology combined with ultra performance liquid chromatography-tandem mass spectrometry.Forage samples were extracted with 8.0 mL of acetonitrile containing 1%formic acid.The extraction mixture was vortexed and then centrifuged at 8 000 r/min for 5 min,after which 1.0 mL of the supernatant was mixed with 3.0 mL of water and vortexed for 0.5 min.The entire mixture was transferred to a solid-phase supported liquid-liquid extraction column and allowed to stand for 10 min.The column was eluted with 6.0 mL of ethyl acetate and the collected eluate was evaporated under nitrogen at temperatures below 40℃.The residue was dissolved in 1 mL of acetonitrile-water(1∶3,volume ratio,containing 0.5%formic acid),and separated using an Acquity UPLC BEH C18 column(100 mm×2.1 mm,1.7 μm).A solution of 0.5%formic acid in water containing 0.25 mmol/L ammonium acetate was used as mobile phase A,while 0.5%formic acid in acetonitrile was used as mobile phase B.The following gradient elution program was used:0-1.0 min,90%B;1.0-5.0 min,90%B-75%B;5.0-10 min,75%B-50%B;10-12 min,50%B-10%B;12-14 min,10%B;14-16 min,10%B-90%B.Ten ergot alkaloid toxins(including three pairs of isomers:ergocornine,ergocristine,ergocristinine,ergocryptine,and ergocryptinine)were effectively separated following gradient elution.Positive electrospray-ionization mode and multiple reaction monitoring(MRM)scanning was used for detection,with an external standard curve used for quantification purposes.The ten ergot alkaloid toxins exhibited good linear rela-tionships within their respective linear ranges(r2>0.995).Alfalfa,Leymus chinensis,oats,and silage corn exhibited LODs of 0.1-2.3 μg/kg for the ten ergot alkaloid toxins,with LOQs of 0.4-7.3 μg/kg.The developed method exhibited overall recovery rates of between 66.3%and 116.7%,with relative standard deviations of less than 9.9%.The matrix effect mainly manifested itself in the form of matrix suppression,with silage corn exhibiting a significantly stronger matrix effect than the other three types of forage.Silage corn exhibited a strong matrix effect in more than 50%of samples,while alfalfa,oats,and Leymus chinensis showed relatively low matrix effects,with more than 63%being weak.The developed method is simple to operate,provides accurate results,and exhibits minimal interference;hence,it is suitable for simultaneously screening and confirming ergot alkaloid toxins in forage,thereby providing technical support for monitoring forage quality,while also expanding the applicability of solid-phase supported liquid-liquid extraction technology in the toxin-detection field.
Ultraviolet(UV)absorbers are a group of chemicals widely used in various industrial and consumer products,such as plastics,coatings,and personal care products,to protect against UV radiation.Among them,benzotriazole derivatives(e.g.UV-326,UV-327,UV-328,UV-329,and UV-P)are the most frequently employed.Owing to their widespread use and potential persistence in the environment,these compounds have been detected in various environmental matrices,including surface water,wastewater,sediment,and biota.Certain UV stabilizers have been reported to exhibit endocrine-disrupting properties and pose potential ecological risks.Therefore,developing sensitive and reliable analytical methods for monitoring these compounds in environmental samples is essential.To address the need for reliable detection methods,this study developed a robust method based on liquid-liquid extraction(LLE)coupled with ultra performance liquid chromatography-triple quadrupole mass spectrometry(UPLC-MS/MS)for the simultaneous de-termination of eight UV absorbers in surface water and wastewater.Critical optimization of the pretreatment process focused on solvent selection and purification parameters.The finalized protocol involved extracting 100 mL water samples twice with dichloromethane.After nitrogen-assisted solvent evaporation,the residue was reconstituted in methanol and mixed with the internal standard so-lution.UPLC-MS/MS parameters were optimized to achieve optimal instrumental performance.The separation of the eight UVs was performed using a BEH C18 column(100 mm×2.1 mm,1.7 μm)with a gradient elution system consisting of 0.2%(mass fraction)formic acid aqueous solution and acetonitrile at a flow rate of 0.4 mL/min.The injection volume was 2 μL.Detection was performed in positive ion mode using multiple reaction monitoring(MRM),with an electrospray ionization voltage set at 5 500 V.Quantification was achieved via internal standard calibration to ensure precision and accuracy.The method demonstrated excellent linearity for all target compounds across their respective concentration ranges,with a correlation coefficient(r)>0.995.The method detection limits(MDLs)ranged from 1.3 ng/L to 2.8 ng/L,indicating high sensitivity.Recovery tests conducted at low,medium,and high spiking levels(20,200,and 800 ng/L)yielded recoveries of 80.3%-117.8%,with relative standard deviations(RSDs)of 1.4%-10.5%,confirming the method's robustness across different sample matrices.Application of the method to 10 textile dyeing wastewater samples revealed the presence of four UV absorbers:UV-329,UV-326,UV-328,and UV-350.Notably,UV-329 showed the highest detection frequency and accounted for 85%of the total detected mass concentrations,ranging from 5.2 to 2 109 ng/L.Its prevalence suggests its widespread use in industrial processes and potential persistence in aquatic environments.In conclusion,the developed method is highly sensitive,accurate,and reliable for detecting UV absorbers in environmental water samples.Its successful application to surface water and wastewater analysis provides a valuable tool for monitoring these emerging contaminants,thereby supporting the assessment of their environmental and health risks.This study highlights the importance of continued monitoring and regulation of UV absorbers to mitigate their potential adverse effects on ecosystems and human health.
Lubaberone （LUB） is the first U S Food and Drug Administration （FDA） approved feed additives for reducing gaseous emissions from animals or their wastes， and the intake of animal-derived foods is an important source of human exposure to LUB. However， the lack of applicable analytical methods makes it difficult for regulatory authorities to monitor LUB in animal-derived foods. There is an urgent need to establish efficient and accurate methods for the analysis of LUB in animal-derived foods. In this study， a method was developed for the determination of LUB from six types of typical animal-derived foods （beef， bovine liver， bovine fat， mutton， sheep liver and sheep fat） by ultra performance liquid chromatography-tandem mass spectrometry （UPLC-MS/MS）. The effects of extraction solvent types， extraction methods， extraction time and purification cartridges on the recovery of LUB were investigated， and the optimal conditions for sample pretreatment were confirmed. The homogenized samples were extracted using 0.5% formic acid acetonitrile via ultrasonication for 10 min， and then filtered by high-speed centrifugation， and the extracted solution was cleaned using SPE with a PRiME HLB cartridge. The LUB was separated on a Shim-pack GIST C18-AQ chromatographic column （100 mm×2.1 mm， 2.7 μm） with 0.1% formic acid aqueous solution-0.1% formic acid acetonitrile as mobile phases at a flow rate of 0.3 mL/min， and detected in positive ion switching mode （ESI+） with multiple reaction monitoring （MRM） scanning， and quantitatively analyzed using the external standard method. Under the optimized experimental conditions， LUB in different animal-derived food matrices showed good linearity within their respective linear concentration ranges with the correlation coefficients （r） greater than 0.99. The limits of detection （LODs） and the limits of quantification （LOQs） were 0.4‒1.0 μg/kg and 1.0‒2.0 μg/kg， respectively. The recoveries of LUB spiked at low， medium and high levels ranged from 81.5% to 116.5% with relative standard deviations （RSDs） of 2.0%‒8.2%. The method is simple， rapid and highly sensitive， which can enable the analysis of LUB residues in animal-derived foods， and provides analytical technology support for the daily detection of LUB residues in imported and exported animal-derived foods.
