Journal of Chromatography A最新文献_第3页

Advances in sample preparation techniques for chromatographic analysis of personal care products and their metabolites in human biological matrices: A critical review 人体生物基质中个人护理产品及其代谢物色谱分析的样品制备技术进展综述

IF 4 2区化学 Q1 BIOCHEMICAL RESEARCH METHODS

Journal of Chromatography A

Pub Date : 2026-04-12 Epub Date: 2026-02-01 DOI: 10.1016/j.chroma.2026.466763

José de Jesús Olmos-Espejel , Jerónimo Cabrera-Peralta , Araceli Peña-Alvarez , Iran Ocaña-Rios

The extensive use of personal care products (PCPs) — including preservatives, UV filters, insect repellents, antimicrobials, and synthetic musks — has led to widespread human exposure, making them contaminants of emerging concern. They are ingredients of products that are used daily for hygiene and skin care. While biomonitoring studies confirm the presence of both parent PCPs and their metabolites in human tissues and fluids, a critical gap exists in the review of analytical methods for analyzing them simultaneously in complex biological matrices. This review critically examines state-of-the-art analytical techniques published from 2013 to the present for the simultaneous determination of multi-class PCPs and their metabolites in human biological samples, such as urine, blood, plasma, serum, saliva, breast milk, nails, and hair. It highlights advances in sample preparation, including innovative extraction and clean-up strategies, as well as chromatographic methods coupled with mass spectrometry. Special emphasis is placed on evaluating key analytical validation parameters, including accuracy, precision, limits of detection, and limits of quantification. Emerging trends reveal that parabens and their metabolites are the most frequently studied compounds, while synthetic musks remain notably underrepresented. Among biological matrices, urine and nails dominate as the preferred liquid and solid samples, respectively. To our knowledge, this is the first review to synthesize recent advances in the simultaneous analysis of diverse PCP classes and their metabolites in human matrices, offering a valuable resource for toxicologists and analytical chemists engaged in exposure assessment and biomonitoring.

个人护理产品（pcp）的广泛使用——包括防腐剂、紫外线过滤器、驱虫剂、抗菌剂和合成麝香——导致了广泛的人类接触，使它们成为日益受到关注的污染物。它们是日常用于卫生和皮肤护理产品的成分。虽然生物监测研究证实母体pcp及其代谢物存在于人体组织和液体中，但在复杂生物基质中同时分析它们的分析方法的审查方面存在一个关键空白。本综述对2013年至今发表的用于同时测定人类生物样品（如尿液、血液、血浆、血清、唾液、母乳、指甲和头发）中多类pcp及其代谢物的最新分析技术进行了严格审查。它强调了样品制备方面的进展，包括创新的提取和清理策略，以及与质谱相结合的色谱方法。特别强调的是评估关键的分析验证参数，包括准确性、精密度、检测限和定量限。新兴趋势表明，对羟基苯甲酸酯及其代谢物是最常被研究的化合物，而合成麝香仍然明显缺乏代表性。在生物基质中，尿液和指甲分别是首选的液体和固体样品。据我们所知，这是第一次综合了人体中多种PCP类别及其代谢物同时分析的最新进展，为从事暴露评估和生物监测的毒理学家和分析化学家提供了宝贵的资源。

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引用次数: 0

A fully automated high-throughput system for simultaneously enriching and separating intact glycopeptides 全自动高通量系统，可同时富集和分离完整的糖肽

IF 4 2区化学 Q1 BIOCHEMICAL RESEARCH METHODS

Journal of Chromatography A

Pub Date : 2026-04-12 Epub Date: 2026-02-04 DOI: 10.1016/j.chroma.2026.466776

Zhonghan Hu , Rong Liu , Junhui Li , Chenlu Wang , Hongxia Wang , Jiancheng Yu , Keqi Tang

An improved fully automated high-throughput system, named as AutoGP 2, for simultaneously enriching and separating intact glycopeptides has been developed in this study by introducing a second 10-port valve (valve 2) to the previously developed AutoGP system with only a 10-port valve (valve 1). Specifically, the addition of the second 10-port valve in the new AutoGP 2 system allows simultaneous enrichment and separation of intact glycopeptides, which significantly improve the sample analysis throughput as compared to previously developed AutoGP system. The optimal operating parameters of the AutoGP 2 were systematically evaluated, and the intra-day and inter-day reproducibility of this system were assessed through 24-h and 15-day long stress tests. The experimental evaluation has shown outstanding sensitivity and reproducibility of AutoGP 2. Over 560 site-specific glycans were identified from 1 μg peptide mixture with median CV of 10.8% in triplicate sample analyses. By coupling the AutoGP 2 with a high resolution Orbitrap mass spectrometer, the duty cycle of the instrument for a single sample analysis was increased from 55% to 80%. Specifically, 166 minutes were saved for every two-sample analyses in the AutoGP 2 as compared with the previous AutoGP system, which increased the sample analysis throughput by 1.5 times. All the experimental results have consistently shown that the AutoGP 2 is well-suited for large cohort clinical glycoproteomic sample analysis and very effective in the sample amount limited glycoproteomic analysis.

本研究开发了一种改进的全自动高通量系统，称为AutoGP 2，用于同时富集和分离完整的糖肽，方法是在之前开发的只有10口阀（阀1）的AutoGP系统上引入第二个10口阀（阀2）。具体来说，在新的AutoGP 2系统中增加了第二个10端口阀，可以同时富集和分离完整的糖肽，与以前开发的AutoGP系统相比，这显着提高了样品分析吞吐量。系统评价了AutoGP 2的最佳操作参数，并通过24小时和15天的压力测试评估了该系统的日内和日内重复性。实验结果表明，AutoGP 2具有良好的灵敏度和重复性。在三次样品分析中，从1个多肽混合物中鉴定出560多个位点特异性聚糖，中位变异系数为10.8%。通过将AutoGP 2与高分辨率Orbitrap质谱仪耦合，仪器对单个样品分析的占空比从55%增加到80%。具体来说，与之前的AutoGP系统相比，AutoGP 2中每两个样品分析节省166分钟，将样品分析吞吐量提高了1.5倍。所有实验结果一致表明，AutoGP 2非常适合于大队列临床糖蛋白组学样本分析，并且在样本量有限的糖蛋白组学分析中非常有效。

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引用次数: 0

The adsorption and enantioselective properties of threonine-based Zn-MOF composite supported on agar biomass aerogel 琼脂生物质气凝胶负载苏氨酸基Zn-MOF复合材料的吸附和对映选择性能

IF 4 2区化学 Q1 BIOCHEMICAL RESEARCH METHODS

Journal of Chromatography A

Pub Date : 2026-04-12 Epub Date: 2026-02-04 DOI: 10.1016/j.chroma.2026.466775

Ksenia S. Balandina , Yan-Rui Wang , Huang-Jie Lu , Natalia A. Osokina , Zhi-Hui Zhang , Vladimir Yu. Guskov

The adsorption activity and enantioselectivity of a novel chiral metal-organic framework based on zinc and a ligand obtained by condensation of threonine and 4-pyridine carbaldehyde (Zn-MOF), as well as its composite with agar (AG/Zn-MOF-0.4), were estimated by inverse gas chromatography. Specific retention volumes (V_g, mL/g) and thermodynamic characteristics of adsorption for different classes of organic substances were calculated from chromatographic experiment. Enantioselectivity was studied with respect to α-pinenes and limonenes, by V_g and adsorption isotherms analysis. A composite material (AG/Zn-MOF-0.4) had more polar surfaces than its parent MOF (Zn-MOF) due to strong interactions with agar. Adsorption analysis showed that the molecules interact with the surface only by specific parts (e.g., a benzene ring or hydroxyl group). Both studied samples also demonstrated enantioselectivity, preferentially adsorbing L-α-pinene. AG/Zn-MOF-0.4 has also shown the ability to recognizing limonene enantiomers. The enantioselectivity coefficients α were higher on AG/Zn-MOF-0.4 than on Zn-MOF or pure agar aerogel. So the combination of chiral MOF with agar created a synergistic effect, enhancing chiral recognition and adsorption activity. This approach is promising for developing advanced chiral adsorbents.

采用反相气相色谱法测定了新型手性金属-有机骨架（Zn-MOF）及其与琼脂（AG/Zn-MOF-0.4）的吸附活性和对映选择性。通过色谱实验计算了不同种类有机物的比保留体积（Vg, mL/g）和吸附热力学特性。通过吸附等温线和Vg分析，研究了其对α-蒎烯和柠檬烯的对映选择性。复合材料（AG/Zn-MOF-0.4）由于与琼脂的强相互作用，具有比母体MOF （Zn-MOF）更多的极性表面。吸附分析表明，分子仅通过特定部分（如苯环或羟基）与表面相互作用。两种样品均表现出对映选择性，优先吸附L-α-蒎烯。AG/Zn-MOF-0.4也显示出识别柠檬烯对映体的能力。AG/Zn-MOF-0.4的对映体选择性系数α高于Zn-MOF或纯琼脂气凝胶。因此，手性MOF与琼脂的结合产生了协同效应，增强了手性识别和吸附活性。该方法有望开发先进的手性吸附剂。

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引用次数: 0

A new HPTLC-image analysis approach for assessing interactions and antioxidant profiles of adrenergic drugs 一种新的hptlc图像分析方法用于评估肾上腺素能药物的相互作用和抗氧化谱

IF 4 2区化学 Q1 BIOCHEMICAL RESEARCH METHODS

Journal of Chromatography A

Pub Date : 2026-04-12 Epub Date: 2026-02-04 DOI: 10.1016/j.chroma.2026.466774

Larisa Silveșan , Claudia Cimpoiu , Dorina Casoni

This study proposes a new approach for the in-vitro evaluation of antioxidant profile of adrenergic drugs using high-performance thin-layer chromatography coupled with image analysis techniques (HPTLC-IA). In this regard, stationary phases of different polarity (Silica gel and RP-8, RP-18 W, CN and DIOL) were selected to simulate different environments and interactions that adrenergic drugs might encounter in the human body. Antioxidant activity was quantified using both DPPH^• and ABTS^⁺• radical scavenging assays on all five stationary phases. Image processing and analysis was used to quantitatively compare the free radical scavenging potential of the analyzed compounds after converting the RGB image into different color channels, namely the green channel for the DPPH test, and the red channel for ABTS. Chromatographic data were processed using multivariate analysis. Results revealed that highly polar compounds, that is norepinephrine, etilefrine, metaraminol and midodrine exhibited strong interactions with silica gel but limited retention on reversed-phase, while lipophilic derivatives, namely naphazoline, xylometazoline, clenbuterol and bambuterol demonstrated strong affinity for non-polar and moderately polar phases, predicting good membrane penetration and blood-brain barrier permeability. Interestingly, some β-agonists (bambuterol, fenoterol, buphenine and irsoxsuprine) with polar groups showed unexpectedly weak silica gel retention, highlighting their dominant lipophilic contributions. Regarding the antioxidant activity, the results showed that phase interactions with the stationary phase significantly influenced the activity of adrenergic drugs. Thus, the polar phases (silica gel, DIOL) enhanced radical scavenging activity, whereas non-polar phases often reduced it. The developed HPTLC-IA method, integrating selective stationary phases and two-radical assays, offers a novel and cost-effective approach for screening the possible variation of antioxidant activity of drugs after their interaction with physiological media constituents.

本研究提出了一种利用高效薄层色谱-图像分析技术（HPTLC-IA）体外评价肾上腺素能药物抗氧化特性的新方法。为此，我们选择了不同极性的固定相（硅胶和RP-8、RP-18 W、CN和DIOL）来模拟肾上腺素能药物在人体内可能遇到的不同环境和相互作用。采用DPPH•和ABTS•对所有5个固定相进行自由基清除实验，定量测定其抗氧化活性。通过图像处理和分析，将RGB图像转换为不同颜色通道，即DPPH测试的绿色通道和ABTS测试的红色通道，定量比较所分析化合物的自由基清除能力。色谱数据采用多变量分析处理。结果表明，高极性化合物（去甲肾上腺素、依替林、甲氨醇和米多宁）与硅胶相互作用强，但在反相保留有限，而亲脂衍生物（萘唑啉、木美唑啉、克仑特罗和班特罗）对非极性和中极性相具有很强的亲和力，预示着良好的膜穿透性和血脑屏障渗透性。有趣的是，一些具有极性基团的β-激动剂（班布特罗、非诺特罗、丁非宁和伊索苏嘌呤）出人意料地表现出较弱的硅胶滞留，突出了它们主要的亲脂作用。在抗氧化活性方面，结果表明，相与固定相的相互作用显著影响肾上腺素能药物的活性。因此，极性相（硅胶，DIOL）增强自由基清除活性，而非极性相通常会降低自由基清除活性。建立的HPTLC-IA方法结合了选择性固定相和双自由基分析，为筛选药物与生理介质成分相互作用后抗氧化活性的可能变化提供了一种新颖而经济的方法。

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引用次数: 0

Modeling retention time distribution in continuous Protein A affinity capture for materials traceability and diversion: A case study with twin-column system 在连续蛋白A亲和捕获中为材料可追溯性和转移建模保留时间分布：双柱系统的案例研究

IF 4 2区化学 Q1 BIOCHEMICAL RESEARCH METHODS

Journal of Chromatography A

Pub Date : 2026-04-12 Epub Date: 2026-02-06 DOI: 10.1016/j.chroma.2026.466780

Wu-Wei Chen , Liang-Zhi Qiao , Alois Jungbauer , Hai-Bin Qu , Shan-Jing Yao , Dong-Qiang Lin

The biopharmaceutical industry is increasingly shifting from batch processing to continuous manufacturing. The ICH Q13 guideline outlines regulatory considerations for the implementation of continuous manufacturing with emphasis on material traceability. Although continuous capture with Protein A affinity chromatography is widely used, the aspect of material traceability remains underexplored. In this study, a retention time distribution (ReTD) model for a twin-column continuous capture process was developed, building on the single column model and the connection condition of the process to enable material traceability. The ReTD behaviors during the startup, interconnected load and disconnected load steps were simulated, focusing on how the material are distributed in the elution peaks across subsequent switches. These simulations were validated through tracer experiments using fluorescently labeled proteins as inert tracer, showing good agreement between the model predictions and experimental results. This observed ReTD behavior is attributed to the combination effects of breakthrough dynamics, exchange effects, and operational step changes. Based on the developed ReTD model, a method for materials traceability and diversion was proposed. The ReTD behavior for materials with extended durations can be extrapolated from materials with duration shorter than one switch, owing to the periodic nature of the continuous capture process. This ReTD-based approach allows for the tracking and evaluation of disturbance entering the system at any time, as well as their persistence for varying durations. These materials can then be diverted according to the acceptable limits, facilitating real-time product release. The proposed ReTD-based method offers a practical solution for ensuring material traceability and diversion, helping meet regulatory requirements and improving real-time product release in continuous capture processes.

生物制药行业正日益从批量生产转向连续生产。ICH Q13指南概述了实施连续生产的法规考虑，重点是材料可追溯性。尽管蛋白A亲和层析的连续捕获被广泛使用，但材料可追溯性方面仍未得到充分探索。本研究建立了双柱连续捕集工艺的保留时间分布（ReTD）模型，以单柱模型和工艺的连接条件为基础，实现了物料的可追溯性。模拟了启动、连接负载和断开负载步骤中的ReTD行为，重点研究了材料如何在随后的开关中分布在洗脱峰中。这些模拟通过示踪实验验证，使用荧光标记的蛋白质作为惰性示踪剂，显示模型预测和实验结果之间的良好一致性。这种观察到的ReTD行为归因于突破动力学、交换效应和操作阶跃变化的综合效应。在建立ReTD模型的基础上，提出了物料溯源和转移的方法。由于连续捕获过程的周期性，具有延长持续时间的材料的ReTD行为可以从持续时间短于一个开关的材料中推断出来。这种基于retd的方法允许在任何时候跟踪和评估进入系统的干扰，以及它们在不同持续时间内的持久性。然后，这些材料可以根据可接受的限制进行转移，从而促进产品的实时释放。提出的基于retd的方法为确保材料的可追溯性和转移提供了一种实用的解决方案，有助于满足监管要求，并改善连续捕获过程中的实时产品释放。

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引用次数: 0

Wood-derived cellulose aerogel decorated with high Zr-MOF loading for simple and efficient extraction of etomidate analogs from urine and e-cigarette samples 高Zr-MOF负载木质纤维素气凝胶用于从尿液和电子烟样品中简单有效地提取依托咪酯类似物。

IF 4 2区化学 Q1 BIOCHEMICAL RESEARCH METHODS

Journal of Chromatography A

Pub Date : 2026-04-12 Epub Date: 2026-02-08 DOI: 10.1016/j.chroma.2026.466783

Mingjie Guo , Weihong Xie , Suling Zhang , Xiaofei Li , Jianwei Zhang , Zhongshi Xu , Ying pan , Dong zhang , Lulu Xu , Ting Lü , Linlin Qiu

Etomidate and its analogs can produce euphoria and addiction, and their recent abuse has led to an increasing number of poisoning and fatal cases. Thus, they are classified as controlled psychotropic drugs. This work proposed small-sized wood aerogel-supported zirconium metal-organic framework (UiO-66) particles for the extraction of trace etomidate and its analogs from urine and e-cigarette samples. Owing to the removal of lignin and hemicellulose, the delignified wood aerogel exhibited enhanced internal pore structures and exposed functional groups, which facilitated the immobilization of more UiO-66 crystals and achieved a high loading efficiency of 61.3 %. The lightweight UiO-66/wood aerogel composite enables facile separation, avoiding UiO-66 powder leakage while retaining its adsorption capacity. The composite was mechanically stable and kept commendable reusability over nine consecutive cycles. Benefiting from its high specific surface area and multiple chemical interactions with target drugs, the extraction method achieved enrichment factors of 22.9–30.0. Key factors influencing extraction—including solution pH, extraction time, the type and volume of desorption solvent, and desorption time—were investigated. Under the optimal conditions, the UiO-66/wood aerogel-based micro-solid phase extraction coupled with GC–MS method showed wide linear ranges and good precision (1.1%–3.2% for intra-day; 3.4%–5.1% for inter-day). The limits of detection and quantification were in the range of 0.02–0.04 μg/L and 0.05–0.1 μg/L, respectively. The recoveries for the urine and e-cigarette oil samples ranged from 86.2%–110.2% and 80.1%–107.9%, respectively. This method is eco-friendly, simple, and sensitive for the determination of new psychotropic drugs in clinical and forensic applications.

依托咪酯及其类似物可以产生欣快感和成瘾，最近滥用它们导致越来越多的中毒和死亡病例。因此，它们被归类为受管制的精神药物。本研究提出了一种小型木质气凝胶支撑的金属锆有机骨架（UiO-66）颗粒，用于从尿液和电子烟样品中提取微量依托咪酯及其类似物。由于木质素和半纤维素的去除，去木质素化木材气凝胶内部孔隙结构增强，官能团暴露，有利于固定化更多的UiO-66晶体，负载效率高达61.3%。轻质UiO-66/木材气凝胶复合材料易于分离，避免UiO-66粉末泄漏，同时保持其吸附能力。该复合材料具有机械稳定性，并在连续9个循环中保持了值得称赞的可重用性。得益于其高比表面积和与目标药物的多重化学相互作用，该提取方法的富集系数为22.9-30.0。考察了影响提取的主要因素：溶液pH、提取时间、解吸溶剂的种类和体积、解吸时间。在最佳条件下，uui -66/木气凝胶微固相萃取- GC-MS法线性范围宽，精密度高（日内1.1% ~ 3.2%，日内3.4% ~ 5.1%）。检测限为0.02 ~ 0.04 μg/L，定量限为0.05 ~ 0.1 μg/L。尿液和电子烟油样品的回收率分别为86.2% ~ 110.2%和80.1% ~ 107.9%。该方法具有生态友好、简便、灵敏等特点，可用于临床和法医学中新型精神药物的测定。

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引用次数: 0

Magnetic C18-modified mesoporous silica as an efficient adsorbent for dispersive solid phase microextraction of cardiac glycosides in human plasma 磁性c18修饰介孔二氧化硅作为分散固相微萃取人血浆中心脏苷的高效吸附剂

IF 4 2区化学 Q1 BIOCHEMICAL RESEARCH METHODS

Journal of Chromatography A

Pub Date : 2026-03-29 Epub Date: 2026-01-23 DOI: 10.1016/j.chroma.2026.466736

Jia Chen , Run-qin Wang , Bei-bei Guo , Li Yang , Shuang Hu

A magnetic mesoporous silica (SBA-15)-based dispersive solid phase microextraction was first proposed and introduced for simultaneous analysis of four cardiac glycosides (including deslanoside, lanatoside C, digoxin, and digitoxin) in human plasma coupled with HPLC-UV. In this study, magnetic C₁₈-modified SBA-15 (SBA-15/C₁₈/Fe₃O₄) was used as the extractant, which was prepared and characterized using a suite of techniques including Fourier transform infrared spectroscopy, transmission electron microscopy, N₂ adsorption-desorption, vibrating sample magnetometer, scanning electron microscopy, energy-dispersive X-ray spectroscopy and X-ray photoelectron spectroscopy. The influencing parameters of the extraction were investigated and optimized. The adsorption process of SBA-15/C₁₈/Fe₃O₄ followed the Langmuir isotherm and pseudo-second-order kinetic models. The adsorbent also exhibited excellent thermal and chemical stability, alongside good reusability. Under the optimum experimental conditions, the proposed method demonstrated satisfactory performance with good linearity (8-3200 ng/mL for deslanoside and 1.6-640 ng/mL for lanatoside C, digoxin and digitoxin with the correlation coefficients no less than 0.9987), high precision (0.8%<RSD<9.7%) and accuracy (-10.3%<RE<8.4%), and acceptable extraction recoveries (56.5%-71.0%). In addition, an absence of matrix effect was observed for the proposed method when applied to human plasma, while satisfactory lower limits of quantification (8 ng/mL for deslanoside and 1.6 ng/mL for lanatoside C, digoxin and digitoxin) and stability were obtained. Overall, owing to its simplicity, efficiency, quickness and eco-friendliness, the developed method serves as a promising alternative for determining cardiac glycosides in complex biological matrices.

首次提出了一种基于磁性介孔二氧化硅（SBA-15）的分散固相微萃取方法，并结合HPLC-UV同时分析了人血浆中的四种心脏苷（包括地黄糖苷、地黄糖苷C、地黄糖苷和地黄糖苷）。本研究以磁性C18改性SBA-15 （SBA-15/C18/Fe3O4）为萃取剂，采用傅里叶变换红外光谱、透射电镜、N2吸附-解吸、振动样品磁强计、扫描电镜、能量色散x射线能谱和x射线光电子能谱等技术对其进行了制备和表征。对影响提取工艺的参数进行了研究和优化。SBA-15/C18/Fe3O4的吸附过程符合Langmuir等温线和拟二级动力学模型。吸附剂还表现出优异的热稳定性和化学稳定性，以及良好的可重复使用性。在最佳实验条件下，该方法具有良好的线性关系（地精苷8 ~ 3200 ng/mL，地精苷C、地高辛和地黄素1.6 ~ 640 ng/mL，相关系数不小于0.9987），精密度（RSD<9.7%）和准确度（-10.3%< rec <8.4%），提取回收率（56.5% ~ 71.0%）。此外，将该方法应用于人血浆时，观察到没有基质效应，同时获得了令人满意的定量下限（地桂苷8 ng/mL，地桂苷C、地高辛和地桂苷1.6 ng/mL）和稳定性。总之，由于其简单、高效、快速和生态友好性，所开发的方法是测定复杂生物基质中心脏苷的有希望的替代方法。

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引用次数: 0

Editorial on the 12th international conference on countercurrent chromatography – CCC2024, Rio de Janeiro, RJ, Brazil 第12届逆流色谱国际会议CCC2024，里约热内卢de Janeiro， RJ， Brazil社论。

IF 4 2区化学 Q1 BIOCHEMICAL RESEARCH METHODS

Journal of Chromatography A

Pub Date : 2026-03-29 Epub Date: 2026-02-05 DOI: 10.1016/j.chroma.2026.466778

Gilda Guimarães Leitão , Martha Knight

引用次数: 0

The role of particle size distribution on permeability and efficiency of slurry-packed liquid chromatography columns 粒径分布对浆料填充柱渗透性和效率的影响

IF 4 2区化学 Q1 BIOCHEMICAL RESEARCH METHODS

Journal of Chromatography A

Pub Date : 2026-03-29 Epub Date: 2026-01-28 DOI: 10.1016/j.chroma.2026.466729

Fabrice Gritti

This review critically examines past and recent studies addressing the intrinsic role of particle size distribution (PSD) in high-performance liquid chromatography (HPLC), with a focus on speed and efficiency. The discussion integrates theoretical models of bed permeability and plate height with experimental evidence to assess the influence of PSD on column permeability and chromatographic resolution. Overall, most findings indicate that PSD has little or no intrinsic impact on key performance metrics, including column permeability, longitudinal diffusion, short-range axial dispersion, and solid–liquid mass transfer, when comparing randomly packed beds of monodisperse and polydisperse particles under equivalent conditions of bed porosity and mean diameters: the Sauter mean diameter for permeability and eddy dispersion and the squared volume-weighted mean particle diameter for slow intraparticle diffusion and high speeds. Rather than reducing particle size polydispersity, efforts should prioritize improved packing methodologies and particle morphologies to minimize long-range structural heterogeneities across the column diameter. While random sphere packings remain the current standard, monodisperse particles may offer some potential for next-generation column technology by enabling highly ordered particulate systems known to deliver superior resolution.

本文回顾了过去和最近的研究，讨论了粒径分布（PSD）在高效液相色谱（HPLC）中的内在作用，重点是速度和效率。讨论结合床层渗透率和板高的理论模型和实验证据来评估PSD对柱渗透率和色谱分辨率的影响。总的来说，大多数研究结果表明，在同等床层孔隙度和平均直径条件下，当比较单分散和多分散颗粒随机填充床时，PSD对关键性能指标（包括柱渗透率、纵向扩散、短距离轴向分散和固液传质）的影响很小或没有内在影响。渗透率和涡流扩散的索特平均直径，以及慢扩散和高速扩散的体积加权平均粒径的平方。与其减少颗粒尺寸的多分散性，不如优先改进填料方法和颗粒形态，以尽量减少跨柱径的长期结构不均匀性。虽然随机球体填料仍然是目前的标准，但单分散颗粒可能为下一代色谱柱技术提供一些潜力，因为它使高度有序的颗粒系统能够提供更高的分辨率。

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引用次数: 0

Analysis of deamination impurity of PS-ONs by chemical derivatization of the enzymatic digest and LC-MS/MS 酶消化化学衍生及LC-MS/MS分析PS-ONs脱胺杂质

IF 4 2区化学 Q1 BIOCHEMICAL RESEARCH METHODS

Journal of Chromatography A

Pub Date : 2026-03-29 Epub Date: 2026-01-26 DOI: 10.1016/j.chroma.2026.466753

Chenyue Zhu , Chuanxiang Yao , Huiting Qin , Wei Zhang , Xinyue Zheng , Siying Zheng , Xinyuan Ye , Ying Xiong , Lili Jiang , Sen Li , Meiqin Wang , Hongliang Jiang , Zhifeng Du

The presence of deamination impurities in phosphorothioate oligonucleotides (PS-ONs) presents a significant challenge to quality control, as their detection is complicated by minimal mass differences from the parent molecule and difficulties in chromatographic separation. Herein, a bottom-up strategy based on enzymatic digestion, chemical derivatization, and liquid chromatography-tandem mass spectrometry (LC-MS/MS) was proposed for the sensitive qualitative and quantitative analysis of trace cytosine deamination in PS-ONs. Fomivirsen (FMVS) and its deamination impurity were used as models. The method involved digestion with S1 nuclease, followed by derivatization of the 5’-phosphorothioate in digests with N-(tert-butyldimethylsilyl)-N-methyltrifluoroacetamide (MTBSTFA) to enhance chromatographic retention and mass spectrometric response. This approach effectively eliminated the need for ion-pairing (IP) reagents. The mass shift upon deamination was amplified in the smaller digested fragments, and a unique isotopic pattern in the derivatized products, specifically, the decrease of the first isotopic peak and the increase of the second isotopic peak was exploited for quantification. The method demonstrated excellent linearity over a weight ratio of deamination product to prototype range from 0.05:1 to 0.9:1. Satisfactory accuracy (80–120%) and precision (RSD < 15%) were achieved. Furthermore, the precise deamination site (C8) within a sample containing 10% deamination impurities was successfully identified. Thus, this method provides a powerful tool for profiling deamination impurities in PS-ONs, with potential applications in quality control, stability studies, and the characterization of other nucleic acid modifications.

硫代寡核苷酸（PS-ONs）中脱胺杂质的存在对质量控制提出了重大挑战，因为它们的检测与母体分子的质量差异很小，并且色谱分离困难。本研究提出了一种基于酶解、化学衍生和液相色谱-串联质谱（LC-MS/MS）的自下而上策略，用于PS-ONs中微量胞嘧啶脱氨的灵敏定性和定量分析。以Fomivirsen （FMVS）及其脱胺杂质为模型。该方法先用S1核酸酶消化，然后用N-（叔丁基二甲基硅基）-N-甲基三氟乙酰胺（MTBSTFA）衍生化消化产物中的5 ' -硫代酸，以提高色谱保留率和质谱响应。这种方法有效地消除了对离子配对（IP）试剂的需求。在较小的消化碎片中，脱氨反应的质量位移被放大，衍生产物中有一种独特的同位素模式，即第一同位素峰的下降和第二同位素峰的增加被用来定量。该方法在脱氨产物与原型的重量比范围为0.05:1至0.9:1之间表现出良好的线性关系。准确度（80-120%）和精密度（RSD < 15%）令人满意。此外，在含有10%脱胺杂质的样品中，成功地鉴定了精确的脱胺位点（C8）。因此，该方法为PS-ONs中脱胺杂质的分析提供了强有力的工具，在质量控制、稳定性研究和其他核酸修饰的表征方面具有潜在的应用前景。

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