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Enhancing Peptide Mapping Sequence Coverage Through an Automated Dual Protease Digest 通过自动双蛋白酶消化增强肽图谱序列覆盖
LCGC Europe Pub Date : 2023-08-01 DOI: 10.56530/lcgc.eu.zq5389j9
Craig Jakes, Silvia Millán-Martín, K. Cook, Dan Bach Kristensen, J. Bones, Sara Carillo
{"title":"Enhancing Peptide Mapping Sequence Coverage Through an Automated Dual Protease Digest","authors":"Craig Jakes, Silvia Millán-Martín, K. Cook, Dan Bach Kristensen, J. Bones, Sara Carillo","doi":"10.56530/lcgc.eu.zq5389j9","DOIUrl":"https://doi.org/10.56530/lcgc.eu.zq5389j9","url":null,"abstract":"Peptide mapping is routinely used in the characterization of monoclonal antibodies (mAbs) for confirmation of the primary sequence and for the detection of post-translational modifications (PTMs). Trypsin is one of the most commonly used proteases in peptide mapping protocols due to its high level of specificity. However, it has been observed that trypsin alone is not always sufficient for full sequence coverage because of the presence of long sequences of hydrophobic amino acids that lack trypsin-specific cleavage sites. In this article, trypsin was combined with chymotrypsin to overcome this loss of sequence coverage. Through the immobilization of these proteases on magnetic beads, and by performing the digestion using an automated platform, a rapid and reproducible digest was achieved with low levels of nonspecific peptides (< 1.3%) and a low number of unique peptides generated across technical replicates (< 6). By using a ratio of 50:50 (v/v) trypsin–chymotrypsin, full sequence coverage was achievable.","PeriodicalId":402085,"journal":{"name":"LCGC Europe","volume":"7 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2023-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"130273888","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Polysorbate Analysis in Biopharmaceutical Applications—A Snapshot of the Analytical Toolbox 聚山梨酸酯分析在生物制药中的应用——分析工具箱的快照
LCGC Europe Pub Date : 2023-08-01 DOI: 10.56530/lcgc.eu.sp6166i2
Piotr Alvarez, Shauni Detremmerie, T. Cucu, G. Vanhoenacker, Steffy Denorme, Cindy Lecluyse, Jorn Deryckere, Bram D’Haenens, F. David, Pat Sandra
{"title":"Polysorbate Analysis in Biopharmaceutical Applications—A Snapshot of the Analytical Toolbox","authors":"Piotr Alvarez, Shauni Detremmerie, T. Cucu, G. Vanhoenacker, Steffy Denorme, Cindy Lecluyse, Jorn Deryckere, Bram D’Haenens, F. David, Pat Sandra","doi":"10.56530/lcgc.eu.sp6166i2","DOIUrl":"https://doi.org/10.56530/lcgc.eu.sp6166i2","url":null,"abstract":"Polysorbates (PS) are added to biopharmaceutical formulations to stabilize proteins and monoclonal antibodies (mAbs) and to prevent aggregation, denaturation, and surface adsorption. Polysorbate analysis can be challenging because of their inherent complexity, polydispersity, and their presence in matrices containing high concentrations of proteins and other excipients such as sugars, amino acids, salts, and buffers. This instalment of “Biopharmaceutical Perspectives” provides an overview of different approaches for the qualitative and quantitative analysis of polysorbates, its impurities and degradants in raw material, intermediate dilutions, drug substance, and drug products. Various gas (GC) and liquid chromatography (LC) methods and accompanying detection techniques are briefly discussed and illustrated with results obtained at the authors’ laboratory.","PeriodicalId":402085,"journal":{"name":"LCGC Europe","volume":"88 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2023-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"126925943","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Functional Copolymers Analysis Using Liquid Chromatography—Sort It Out 功能共聚物的液相色谱分析-分选
LCGC Europe Pub Date : 2023-06-01 DOI: 10.56530/lcgc.eu.ku3485u9
Ron A. H. Peters
{"title":"Functional Copolymers Analysis Using Liquid Chromatography—Sort It Out","authors":"Ron A. H. Peters","doi":"10.56530/lcgc.eu.ku3485u9","DOIUrl":"https://doi.org/10.56530/lcgc.eu.ku3485u9","url":null,"abstract":"The latest developments and challenges of using liquid chromatography (LC) for copolymer analysis are discussed.","PeriodicalId":402085,"journal":{"name":"LCGC Europe","volume":"332 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2023-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"133934257","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Approaches to Accelerate Liquid Chromatography Method Development in the Laboratory Using Chemometrics and Machine Learning
LCGC Europe Pub Date : 2023-06-01 DOI: 10.56530/lcgc.eu.rh7676j5
Gerben V. van Henten, Tijmen S Bos, B. Pirok
{"title":"Approaches to Accelerate Liquid Chromatography Method Development in the Laboratory Using Chemometrics and Machine Learning","authors":"Gerben V. van Henten, Tijmen S Bos, B. Pirok","doi":"10.56530/lcgc.eu.rh7676j5","DOIUrl":"https://doi.org/10.56530/lcgc.eu.rh7676j5","url":null,"abstract":"This review examines different workflows that have been designed and used to facilitate and/or automate method development in liquid chromatography (LC).","PeriodicalId":402085,"journal":{"name":"LCGC Europe","volume":"22 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2023-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"124481924","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 2
Micro-Pillar Array Column Separations for Proteomics of Liver Organoids 肝类器官蛋白质组学的微柱阵列柱分离
LCGC Europe Pub Date : 2023-05-02 DOI: 10.56530/lcgc.eu.st2089i6
A. Aizenshtadt, Lise Midtøy, B. Thiede, Stefan Krauss, H. Røberg-Larsen, S. Wilson
{"title":"Micro-Pillar Array Column Separations for Proteomics of Liver Organoids","authors":"A. Aizenshtadt, Lise Midtøy, B. Thiede, Stefan Krauss, H. Røberg-Larsen, S. Wilson","doi":"10.56530/lcgc.eu.st2089i6","DOIUrl":"https://doi.org/10.56530/lcgc.eu.st2089i6","url":null,"abstract":"Organoids are laboratory-grown three-dimensional (3D) models of organs and are emerging tools for studies into developmental biology, drug discovery, and toxicology. Organoids are complex biological materials, and proteomics studies of organoids can benefit from applying high-resolution chromatography devices before mass spectrometric analysis. Micro-pillar array columns have been shown to provide excellent resolution of peptide products of proteolytic digestion of proteins for bottom-up proteomics.This article describes a workflow incorporating a micro-pillar array column for mapping the proteome of human stem cell-derived liver organoids (sample preparation using a SPEED protocol) using trapped ion mobility time-of-flight mass spectrometry (timsTOF-MS).","PeriodicalId":402085,"journal":{"name":"LCGC Europe","volume":"11 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2023-05-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"127445139","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
When van Deemter Goes Upside Down in Chiral Chromatography 当van Deemter在手性色谱中颠倒过来时
LCGC Europe Pub Date : 2023-05-02 DOI: 10.56530/lcgc.eu.st5666h2
Martina Catania, A. Cavazzini, C. De Luca, Simona Felletti
{"title":"When van Deemter Goes Upside Down in Chiral Chromatography","authors":"Martina Catania, A. Cavazzini, C. De Luca, Simona Felletti","doi":"10.56530/lcgc.eu.st5666h2","DOIUrl":"https://doi.org/10.56530/lcgc.eu.st5666h2","url":null,"abstract":"The occurrence of convex-upward van Deemter curves is rare but not unusual in chiral liquid chromatography (LC). In this article, this behaviour, experimentally observed for the more retained enantiomer of a chiral sulfoxide on a polysaccharide-based chiral stationary phase (CSP), has been investigated from an experimental viewpoint, finding that this species is characterized by a very strong, localized adsorption on the CSP. The observation of the behaviour of other chiral sulfoxides with different chemical structures has suggested correlations between molecular properties and specific interactions on the CSP.","PeriodicalId":402085,"journal":{"name":"LCGC Europe","volume":"44 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2023-05-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"128990635","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Evaluation of PGC Stationary Phases Under High-Temperature LC Conditions for the Analysis of Parabens in Food Samples 高温LC条件下PGC固定相分析食品样品中对羟基苯甲酸酯的评价
LCGC Europe Pub Date : 2023-05-02 DOI: 10.56530/lcgc.eu.ng3276e1
Roberta La Tella, Francesca Rigano, Clinton R. Corman, Gabriel A. Odugbesi, P. Donato, P. Dugo, L. Mondello
{"title":"Evaluation of PGC Stationary Phases Under High-Temperature LC Conditions for the Analysis of Parabens in Food Samples","authors":"Roberta La Tella, Francesca Rigano, Clinton R. Corman, Gabriel A. Odugbesi, P. Donato, P. Dugo, L. Mondello","doi":"10.56530/lcgc.eu.ng3276e1","DOIUrl":"https://doi.org/10.56530/lcgc.eu.ng3276e1","url":null,"abstract":"Porous graphitic carbon (PGC) columns for liquid chromatography (LC) represent an alternative to octadecyl‑bonded silica columns for the separation of both polar and nonpolar molecules. This is accomplished by exploiting the polarizability of the stationary phase interacting with the functional groups of the analytes. However, the elution of nonpolar compounds requires a high percentage of organic solvent, losing the intrinsic advantage of reversed‑phase aqueous separations. In this article, we aimed to exploit an additional advantage of such columns, viz. the resistance at high temperatures. Superheated water was employed as the mobile phase, taking advantage of the decrease in water dielectric constant by increasing the temperature. In this context, our goal was to minimize the percentage of organic solvent utilizing high temperatures (up to 250 °C) to achieve fast and “green” separations. The new developed high-temperature LC instrument was applied to the analysis of parabens in food samples.","PeriodicalId":402085,"journal":{"name":"LCGC Europe","volume":"5 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2023-05-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"132677599","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 1
Evaluation of Kinetic Performance of Reversed-Phase Columns for Protein Separations by Gradient Kinetic Plots 用梯度动力学图评价反相色谱柱分离蛋白质的动力学性能
LCGC Europe Pub Date : 2023-05-02 DOI: 10.56530/lcgc.eu.bm1469g6
Simon Jaag, M. Lämmerhofer
{"title":"Evaluation of Kinetic Performance of Reversed-Phase Columns for Protein Separations by Gradient Kinetic Plots","authors":"Simon Jaag, M. Lämmerhofer","doi":"10.56530/lcgc.eu.bm1469g6","DOIUrl":"https://doi.org/10.56530/lcgc.eu.bm1469g6","url":null,"abstract":"The increasing importance of protein biopharmaceuticals has triggered the development of new, highly efficient stationary phases for reversed-phase liquid chromatography (LC) of proteins. They typically have C4 ligands or phenyl surfaces for weak hydrophobic interactions and are based on various morphologies, such as silica monolith, sub-2-µm fully porous particles (FPPs), or superficially porous particles (SPPs). Selection of the best column based on physical parameters provided by vendors may sometimes be unequivocal. Simple performance evaluation tools, such as the gradient kinetic plot methodology reported by Desmet and co-workers, can help to shed light on this issue and allow for a more differentiated view on column performance. This article compares the performance of wide-pore silica monolithic, sub-2-µm FPP, and SPP columns. It also addresses the question of whether 1000 Å or 400 Å SPP columns are more suitable for reversed-phase LC-type protein separations and presents a kinetic performance comparison of a number of wide-pore core–shell particle columns.","PeriodicalId":402085,"journal":{"name":"LCGC Europe","volume":"278 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2023-05-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"114481787","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Monitoring Contaminants of Emerging Concern in the Environment 监测环境中新出现的污染物
LCGC Europe Pub Date : 2023-05-01 DOI: 10.56530/lcgc.eu.cg2286r3
L. Barron
{"title":"Monitoring Contaminants of Emerging Concern in the Environment","authors":"L. Barron","doi":"10.56530/lcgc.eu.cg2286r3","DOIUrl":"https://doi.org/10.56530/lcgc.eu.cg2286r3","url":null,"abstract":"LCGC Europe spoke to Leon Barron from Imperial College London, UK, about his work developing a fully miniaturized workflow incorporating a three-dimensional-printed passive sampler device (3D-PSD) to monitor contaminants of emerging concern (CECs) in river water.","PeriodicalId":402085,"journal":{"name":"LCGC Europe","volume":"210 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2023-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"123389146","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Suspect Screening of Lupin-Produced Phytotoxins in Environmental Samples 环境样品中羽扇豆素产生的植物毒素的可疑筛选
LCGC Europe Pub Date : 2023-05-01 DOI: 10.56530/lcgc.eu.qf7583j1
Xiao-Xing Liang, J. H. Christensen, T. Bucheli, N. J. Nielsen
{"title":"Suspect Screening of Lupin-Produced Phytotoxins in Environmental Samples","authors":"Xiao-Xing Liang, J. H. Christensen, T. Bucheli, N. J. Nielsen","doi":"10.56530/lcgc.eu.qf7583j1","DOIUrl":"https://doi.org/10.56530/lcgc.eu.qf7583j1","url":null,"abstract":"Phytotoxins, the secondary metabolites synthesized by plants, are now recognized as a new category of environmental micropollutants. So far, only a limited number of phytotoxins have been detected and reported in terrestrial and aquatic environments, partially due to the analytical challenge involved. In this article, a robust, reliable, and efficient approach, namely Source Supported Suspect Screening (4S), is introduced for high-throughput analysis of phytotoxins from their plant origin to the downstream environmental compartments. The approach was established on an ultrahigh-performance liquid chromatography–electrospray ionization–quadrupole‑time‑of‑flight-mass spectrometry (UHPLC–ESI-QTOF-MS) analytical platform where optimal sample preparation and chromatographic conditions were investigated and a new suspect screening protocol was suggested.","PeriodicalId":402085,"journal":{"name":"LCGC Europe","volume":"30 1","pages":"0"},"PeriodicalIF":0.0,"publicationDate":"2023-05-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"125515817","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
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