Analytical Chemistry最新文献

{"title":"Task-Dependent Synergy between FTIR and Raman Spectroscopy for Sperm Quality Prediction","authors":"Beatrice Campanella,&nbsp;Lorenzo Birindelli,&nbsp;Stefano Legnaioli,&nbsp;Giulia Lorenzetti,&nbsp;Cristina Consani,&nbsp;Valeria Ales,&nbsp;Edoardo Benedetti,&nbsp;Maria Antonella Bertozzi,&nbsp;Emilia Bramanti","doi":"10.1021/acs.analchem.6c04960","DOIUrl":"10.1021/acs.analchem.6c04960","url":null,"abstract":"<p>Maleinfertility affects approximately 15% of couples worldwide,contributing to 40–50% of infertility cases in Western countries.Standard semen analysis remains the cornerstone of fertility assessment,yet conventional parameters (concentration, motility, morphology)have limited predictive value for fertilization success. Vibrationalspectroscopy, particularly Fourier transform infrared (FTIR) and Ramanspectroscopy, offers rapid, nondestructive chemical fingerprintingof biological samples. However, optimal strategies for integratingmultimodal spectroscopic data remain unclear, and the value of fusionmay depend on the biological end point. This study systematicallycompares FTIR and Raman spectroscopy, both independently and throughmultiple fusion strategies, for predicting rectilinear progressivesperm motility (binary classification) and normal morphology (continuousregression) in human seminal plasma. We determine whether complementaryinformation exists, when fusion provides value, and which strategyoptimally leverages multimodal data. Building on Raman-derived MultivariateCurve Resolution-Alternating Least Squares (MCR-ALS) concentrationprofiles, we further develop a clinical classifier based on four seminalplasma biomarkers (human serum albumin, glutathione S-transferase,transferrin, lysophosphatidylcholine), validated by leave-one-patient-outcross-validation and Firth’s penalized logistic regressionto address near-complete separation in this small-sample setting (n= 30). Our findings reveal technique-specific advantages: Raman spectroscopysubstantially outperforms FTIR for binary motility classification,with feature-level fusion matching but not exceeding Raman’sperformance, indicating limited complementarity. Conversely, FTIRalone achieves superior predictive accuracy for continuous morphologicaloutcomes. The clinical classifier based on MCR-ALS-derived biomarkersachieved AUC (Area Under the ROC Curve) = 0.855, accuracy = 86.7%,and BER (Balanced Error Rate) = 0.136, with model significance confirmedby permutation testing. Elevated human serum albumin and lysophosphatidylcholineemerged as the strongest negative predictors of high motility, consistentwith seminal tract inflammation and sperm-cytotoxic phospholipid accumulation,while transferrin showed a positive association in line with its rolein iron availability for sperm function. These results demonstratethat optimal spectroscopic approaches depend on the biological endpoint, with Raman’s structural sensitivity favoring motilityassessment and FTIR’s compositional profiling better capturingmorphological variation. The biomarker-based classifier is easierto interpret and complements whole-spectrum models by bridging chemometricanalysis and biological interpretation.</p>","PeriodicalId":27,"journal":{"name":"Analytical Chemistry","volume":"98 41","pages":"29684–29699"},"PeriodicalIF":7.3,"publicationDate":"2026-10-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148973812","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"p-Type Dopant-Mediated Electron–Hole Recombineering to Tune the Electrochemiluminescence of Metal–Organic Frameworks","authors":"Guijun Li,&nbsp;Shuai Luo,&nbsp;Si Zhang,&nbsp;Mengjiao Li,&nbsp;Huipu Liu,&nbsp;Huangxian Ju","doi":"10.1021/acs.analchem.6c03746","DOIUrl":"10.1021/acs.analchem.6c03746","url":null,"abstract":"<p>Constructing high-performance electrochemiluminescence(ECL) emittersthrough rational structural engineering is pivotal to overcoming theintrinsic limitations of conventional systems and advancing theirpractical applications. This work reports an electron–holerecombination engineering strategy to enhance the ECL emission ofmetal–organic frameworks (MOFs) by p-type doping of metal single-atomsites. Using Zr-TBAPy as a model, the experimental data and theoreticalcalculations demonstrate that the doping of single-atom Au inducesnotable defect energy levels, thereby providing additional recombinationchannels for conduction-band (CB) electrons and valence-band (VB)holes. The defect level ultimately increases the electron–holerecombination efficiency. Compared to Zr-TBAPy, single-atom Au-dopedZr-TBAPy (Au@Zr-TBAPy) exhibits a 4-fold enhancement in cathodic ECLintensity with an ECL efficiency of up to 50.56%. An ECL immunoassaywas used to demonstrate the ECL application of Au@Zr-TBAPy. This worknot only proposes a rational metal single-atom doping strategy forengineering ECL emitters to achieve high performance but also providesfundamental insights into the dependence of ECL emission on metalsingle-atom sites, opening new avenues for advanced applications ofMOFs in sensitive ECL biosensing.</p>","PeriodicalId":27,"journal":{"name":"Analytical Chemistry","volume":"98 41","pages":"29579–29587"},"PeriodicalIF":7.3,"publicationDate":"2026-10-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149019588","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Real-Time Monitoring Ferroptosis-Mediated Drug-Induced Liver Injury Using a Near-Infrared Mitochondrial Viscosity Probe","authors":"Ben Liu,&nbsp;Yiwu Qiu,&nbsp;Long He,&nbsp;Yi Li","doi":"10.1021/acs.analchem.6c04129","DOIUrl":"10.1021/acs.analchem.6c04129","url":null,"abstract":"<p>Drug-induced liver injury (DILI) is a frequent clinicalconditionin which hepatocyte ferroptosis serves as a core pathological mechanism,leading to critical alterations in the mitochondrial microenvironment,including viscosity. However, visualizing mitochondrial viscositychanges in real time during ferroptosis-driven DILI remains technicallychallenging. Herein, we report XT-TPE, a mitochondrion-targeted near-infrared(NIR) “turn-on” viscosity probe based on a tetraphenylethylene(TPE)-xanthene hybrid. This probe gives a remarkable 129-fold fluorescenceincrease in high-viscosity media with an outstanding linear relationship(<em>R</em>² &gt; 0.999). The probe displays lowcytotoxicitytoward hepatocytes, excellent mitochondrial colocalization (Pearson’scoefficient: 0.916), and enables sensitive monitoring of viscosityfluctuations during ferroptosis-mediated DILI both <em>in vitro</em> and <em>in vivo</em>. Using this probe, we screened 14 redox-activecompounds and identified N-acetylcysteine (NAC) and ferrostatin-1(Fer-1) as strong inhibitors of ferroptosis-driven DILI. In acetaminophen-inducedDILI mouse models, XT-TPE enables noninvasive hepatic viscosity imagingand confirms the therapeutic efficacy of NAC and Fer-1. This workprovides a reliable molecular tool for ferroptosis-related viscosityimaging and establishes a new strategy for anti-DILI drug screening.</p>","PeriodicalId":27,"journal":{"name":"Analytical Chemistry","volume":"98 41","pages":"29616–29626"},"PeriodicalIF":7.3,"publicationDate":"2026-10-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149049986","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Quantification of Intact MDa Poly(A)-Tailed and Non-Tailed mRNA Fractions with Charge Detection Mass Spectrometry","authors":"Julian Robles,&nbsp;Zachary M. Miller,&nbsp;Aaron M. Roe,&nbsp;Stephanie M. Thibert,&nbsp;Matthew R. Gardner,&nbsp;Janelle Tricia M. Magaling,&nbsp;Justin W. Torpey,&nbsp;Angelica F. Castañeda,&nbsp;Evan R. Williams","doi":"10.1021/acs.analchem.6c03007","DOIUrl":"10.1021/acs.analchem.6c03007","url":null,"abstract":"<p>The clinical success of messenger ribonucleic acid (mRNA)therapeuticshas driven the need for robust analytical techniques capable of characterizingmixtures of megadalton (MDa) constructs. A critical attribute of thesetherapeutics is the presence and length of the 3′ poly(A) tail,which governs mRNA stability and translational efficiency. Chargedetection mass spectrometry (CDMS) was used to distinguish and quantifyintact MDa mRNA variants differing only by the poly(A) content. Initialanalysis under native-like conditions was limited by salt adduction,but a 75% methanol solution resulted in nearly a five-fold increasein charge (from ∼60<em>e</em> to ∼280<em>e</em>) and substantially less adduction. Measured masses ofthe intact mRNA were 1.054 MDa and 1.027 MDa for the tailed and non-tailedmRNA constructs, respectively, consistent with the theoretical 26.6kDa mass difference. Mixtures of various compositions resulted inbroad, unresolved peaks at intermediate mass owing to residual saltadduction. A constrained Gaussian deconvolution of these data enabledaccurate quantification across seven different mole fractions (<em>R</em>² = 0.978). Notably, an analysis of a subpopulationof the highest charge-state ions revealed separation into two distinctpeaks, demonstrating that mass resolution for MDa mRNA analysis islimited by adduction rather than instrumental factors, motivatingadditional studies on methods to increase charging for more accuratequantification. These results establish CDMS as a powerful, top-downmethod for precise monitoring of mRNA heterogeneity and degradationin high molecular weight therapeutics.</p>","PeriodicalId":27,"journal":{"name":"Analytical Chemistry","volume":"98 41","pages":"29507–29514"},"PeriodicalIF":7.3,"publicationDate":"2026-10-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149091910","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Multiway Modeling of 2D NMR Spectra for Enhanced Chemical Characterization of Complex Mixtures","authors":"Pia S. Mayer,&nbsp;Søren B. Engelsen,&nbsp;Jesper L. Hinrich,&nbsp;Morten Mørup,&nbsp;Bekzod Khakimov","doi":"10.1021/acs.analchem.6c04147","DOIUrl":"10.1021/acs.analchem.6c04147","url":null,"abstract":"<p>One-dimensional(1D) proton nuclear magnetic resonance (NMR) spectroscopyis widely used for complex mixture analysis because of its high reproducibility,minimal sample preparation, and inherent quantitative capabilities.However, 1D NMR remains constrained by three interrelated limitations:signal overlap among compounds, chemical-shift variability of thesame compounds across samples, and masking of low signal-to-noisefeatures by neighboring high-intensity resonances. Two-dimensional(2D) NMR spectroscopy can reduce these limitations by dispersing spectralinformation over a second dimension and providing additional structuralinformation. Yet its broader implementation in large-scale studiesremains limited by a major data-processing bottleneck: the analysisof multisample, high-dimensional 2D NMR data sets. When acquired acrossmultiple samples, 2D NMR spectra form three-way tensors that containricher chemical information than conventional 1D data sets but arenot readily handled by standard NMR data analysis workflows. In thispaper, we discuss how localized 2D spectral alignment combined withtensor decomposition and shift-tolerant multilinear modeling, includingPARAFAC, PARAFAC2, and shift-invariant approaches, can recover overlapped,shifted, and low-intensity signals from complex 2D NMR spectra. Theseworkflows hold considerable promise for advancing NMR-based complexmixture analysis by expanding metabolite coverage, improving deconvolutionof elusive signals, and enabling more reliable quantification of low-abundancebiomarkers in large-scale studies.</p>","PeriodicalId":27,"journal":{"name":"Analytical Chemistry","volume":"98 41","pages":"29417–29432"},"PeriodicalIF":7.3,"publicationDate":"2026-10-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149091912","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Genetically Encoded APEX2 Enables Metal-Enhanced X-ray Contrast for Energy-Selective Imaging in Complex Tissues","authors":"Dapeng Yin,&nbsp;Qiaowei Tang,&nbsp;Yiliu Wang,&nbsp;Huating Kong,&nbsp;Yong Guan,&nbsp;Ying Zhu","doi":"10.1021/acs.analchem.6c03949","DOIUrl":"https://doi.org/10.1021/acs.analchem.6c03949","url":null,"abstract":"<p>X-ray microscopy offers deep tissuepenetration and highspatialresolution, but its application to genetically targeted imaging remainslimited by the lack of probes capable of generating sufficient andselective X-ray contrast in complex biological tissues. Here we reporta genetically encoded X-ray contrast generation strategy that combinesAAV-mediated expression of engineered peroxidase APEX2, enzymaticsignal amplification through diaminobenzidine (DAB) deposition, andenergy-selective synchrotron X-ray imaging. Optimization of Cre-dependentAAV expression and tissue fixation enabled efficient conversion ofgene expression into localized X-ray-absorbing deposits. We furtheridentified tissue-background absorption as a major factor limitingthe analytical performance of conventional DAB-based X-ray imagingin dense brain regions. To address this challenge, nickel-enhancedDAB (EDAB) was introduced to generate metal-containing reaction productswith characteristic absorption properties. Imaging near the Ni L-edgesubstantially improved signal discrimination between reporter-labeledneurons and surrounding tissue, overcoming the contrast limitationsobserved with conventional DAB imaging. This work establishes a geneticallyencoded and energy-selective X-ray contrast-generation platform thatintegrates reporter engineering, signal amplification, and absorption-edge-guideddetection, providing a general analytical framework for targeted X-rayimaging in complex biological systems.</p>","PeriodicalId":27,"journal":{"name":"Analytical Chemistry","volume":"98 41","pages":"29596–29603"},"PeriodicalIF":7.3,"publicationDate":"2026-10-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149218979","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Ratio-Consistency-Driven Discrimination of Structurally Similar VOCs in PTR-MS","authors":"Ke Wang,Yihao Wang,Xuejiao Huang,Leizi Jiao,Chunjiang Zhao,Daming Dong","doi":"10.1021/acs.analchem.6c02674","DOIUrl":"https://doi.org/10.1021/acs.analchem.6c02674","url":null,"abstract":"Plant volatile organic compounds (VOCs) provide important chemical information related to plant physiological states and plant−environment interactions. Proton transfer reaction mass spectrometry (PTR-MS) enables real-time VOC monitoring with high sensitivity, but overlapping protonated and fragment-ion signals limit the discrimination of structural isomers and mass-similar VOCs. This study proposes a ratio-consistency framework for VOC discrimination under fixed E/N conditions. Centered log-ratio (CLR) transformation is used to represent relative fragment-ion abundance relationships, while a ratio-consistency constraint guides the model to exploit reproducible fragmentation-related patterns and reduce its reliance on absolute intensity variations. For eight structurally similar VOCs, the proposed framework improves closed-set classification accuracy from approximately 55% using intensity-based features to 89.2%. An open-set recognition strategy combining maximum Softmax confidence and ratio-consistency deviation is further introduced, achieving an overall recognition accuracy of 81.1% across known and unknown samples. The framework is subsequently applied to VOC emissions from mechanically wounded tomato plants, where PTR-MS analysis identifies β-pinene as a major VOC released during the early stage of wounding, with independent FTIR measurements providing consistent spectroscopic support. These results demonstrate that relative fragment-ion abundance patterns provide complementary discriminative information for PTR-MS analysis and offer a practical strategy for improving the discrimination and open-set recognition of structurally similar VOCs without additional instrumental modulation.","PeriodicalId":27,"journal":{"name":"Analytical Chemistry","volume":"11 1","pages":""},"PeriodicalIF":7.4,"publicationDate":"2026-10-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149218841","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Second-Derivative IR Spectroscopy Reveals Hidden Modes of Structural Organization within Conformational Disorder","authors":"Diletta Ami,Jacopo Vertemara,Marco Mangiagalli,Paolo Mereghetti,Mattia Pietro Fossati,Greta Bianchi,Simone Vesentini,Stefania Brocca,Giuseppe Zampella,Antonino Natalello","doi":"10.1021/acs.analchem.6c04419","DOIUrl":"https://doi.org/10.1021/acs.analchem.6c04419","url":null,"abstract":"Intrinsically disordered proteins (IDPs) and regions constitute a significant fraction of the proteome. Unlike globular proteins, IDPs exist as dynamic conformational ensembles, allowing functional versatility while challenging conventional structural methods, necessitating an integrated, multi-method approach. Here, we systematically compare IDPs and globular proteins using Fourier Transform Infrared (FTIR) and circular dichroism (CD) spectroscopies to investigate not simply the presence of disorder but also its structural organization. By analyzing the second derivatives of the Amide I′ absorption spectra, we identified a distinct spectral fingerprint of IDPs. Detailed analysis reveals that the spectral features encode distinct modes of structural organization within conformationally heterogeneous ensembles. Notably, the integration of second-derivative FTIR (SD-IR) and CD spectroscopies uncovers conformational states that are not captured by conventional techniques devoted to secondary structure characterization. Specifically, we identified conditions in which proteins retain a conformational order despite exhibiting random coil CD signatures (“ordered random coils”) as well as states displaying canonical α-helical CD features while maintaining substantial structural heterogeneity (“disordered α-helices”). Molecular dynamics simulations support these assignments, revealing the structural basis underlying the spectroscopic findings. Finally, we demonstrate the efficacy of SD-IR spectroscopy for validating putative IDPs, exemplified by the silkworm protein sericin. Overall, this study establishes second-derivative analysis of the Amide I′ band not merely as a diagnostic marker of disorder but also as a powerful approach to resolve distinct organizational states within conformationally heterogeneous protein ensembles.","PeriodicalId":27,"journal":{"name":"Analytical Chemistry","volume":"5 1","pages":""},"PeriodicalIF":7.4,"publicationDate":"2026-10-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149218836","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"DNA-Mediated Self-Assembled SERS Platform Combined with Machine Learning for Therapeutic Drug Monitoring","authors":"Ruiyao Ma,Dan Li,Haoyu Ji,Yaowen Xing,Qiushuang Liu,Zhidan Sun,Yujiang Huang,Ling Xia,Shen Jiang,Xinyue Hu,Lixin Guo,Yang Li","doi":"10.1021/acs.analchem.6c05327","DOIUrl":"https://doi.org/10.1021/acs.analchem.6c05327","url":null,"abstract":"Therapeutic drug monitoring (TDM) critically requires rapid, sensitive, and practical methods for personalized medicine. Herein, we present a label-free surface-enhanced Raman spectroscopy (SERS) platform based on self-assembly of T12 DNA on Ag@ITCNPs mediated by Ca2+. This substrate generates abundant electromagnetic hotspots via electrostatic crosslinking and utilizes the intrinsic thymine peak as an internal standard to effectively eliminate serum matrix interference, achieving an analytical enhancement factor (AEF) of 5.93 × 1013. The platform enables ultrasensitive detection of various antiepileptics and immunosuppressants (e.g., cyclosporine A, carbamazepine, valproate) at the fg/mL level, with excellent reproducibility (RSD = 4.10%). By integrating unsupervised algorithms (PCA, t-SNE) and supervised models (CNN, SVM, KNN, LDA, RF, RNN, XGBoost), it achieves near 100% accuracy in discriminating between different drugs and even among individual patients receiving the same medication. Furthermore, regression models based on SERS enable precise quantification, with results showing excellent agreement with the LC−MS/MS gold standard. This work demonstrates that the synergy between self-assembly directed by DNA and machine learning provides a powerful strategy for constructing highly sensitive and selective SERS platforms, offering a promising solution for rapid therapeutic drug monitoring at the point of care in complex biological matrices.","PeriodicalId":27,"journal":{"name":"Analytical Chemistry","volume":"7 1","pages":""},"PeriodicalIF":7.4,"publicationDate":"2026-10-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149218840","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Automated Quenching and Extraction for High-Precision Metabolomics Uncovers Distinct Metabolome Trajectories across Escherichia coli Lineages","authors":"Kenya Tanaka,Takahiro Bamba,Musashi Takenaka,Yoshimi Hori,Takanobu Yoshida,Mami Matsuda,Akihiko Kondo,Yoshihiro Hayakawa,Tomohisa Hasunuma","doi":"10.1021/acs.analchem.6c01820","DOIUrl":"https://doi.org/10.1021/acs.analchem.6c01820","url":null,"abstract":"Quantitative microbial metabolomics requires rapid quenching and highly standardized extraction, yet these pre-analytical steps remain labor-intensive and difficult to reproduce, limiting time-resolved intracellular measurements under controlled bioreactor cultivation. Here, we present an integrated robotic platform that couples parallel fermenter operation with circulation-based sampling, immediate cold-alcohol quenching, automated refrigerated centrifugation, interface-aware supernatant removal, and solvent extraction with controlled heating and mixing, followed by plate-based storage for liquid chromatography–tandem mass spectrometry (LC–MS/MS). To benchmark the precision of the automated preparation module, we compared repeated LC–MS/MS measurements of a single extract with measurements of independently processed samples subjected to post-quench automated harvesting, extraction, and transfer. The analysis was restricted to 76 analytes consistently detected in all replicates of both datasets. The mean coefficients of variation increased from 6.92% for repeated LC–MS/MS measurements to 8.82% for automated preparation, corresponding to a modest increase of 1.90 percentage points. These results indicate that post-quench automated preparation introduces a small but measurable additional source of variability while maintaining mean analytical variation below 10%. We then performed time-course endo- and exometabolomics across four widely used Escherichia coli lineages (BW25113, BL21(DE3), W, and C). Principal component analysis resolved strain-specific trajectories throughout batch growth and revealed strain-specific metabolic phenotypes, including coordinated accumulation of sugar phosphates and 2-C-methyl-d-erythritol 4-phosphate (MEP) pathway intermediates in strain C, and comparatively high intracellular pools of branched-chain amino acids in strain W. This platform enables standardized and traceable automated preparation for dense, time-resolved metabolomics and provides directly measurable phenotypes for host selection and metabolic engineering.","PeriodicalId":27,"journal":{"name":"Analytical Chemistry","volume":"73 1","pages":""},"PeriodicalIF":7.4,"publicationDate":"2026-10-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"149210895","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}