Investigation of sample handling steps for accurate heparan sulphate disaccharide analysis using HPLC-MS.

IF 3 3区生物学 Q2 BIOCHEMICAL RESEARCH METHODS

ELECTROPHORESIS Pub Date : 2024-08-21 DOI:10.1002/elps.202400091

Domonkos Pál, Gábor Tóth, Lilla Turiák

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引用次数: 0

Abstract

Heparan sulphates (HSs), a specific class of glycosaminoglycans (GAGs), are important participants of cellular signalling. Analytical characterization of GAGs requires a complex sample preparation workflow. Although a detailed stability and recovery study is available for the chondroitin sulphate GAG class, the literature concerning HS is incomplete in this regard. Therefore, our aim was to systematically investigate various parameters that could potentially influence the stability and recovery of HS samples when performing disaccharide analysis using high-performance liquid chromatography-mass spectrometry. First, effects concerning vacuum evaporation and freezing were investigated. Next, the storage stability of the HS disaccharides was analysed under several conditions such as temperature, pH, digestion buffers, injection solvents and storage vessels. We have identified several critical parameters influencing the stability and recovery of HS disaccharides. We concluded that major sample loss is expected when Tris-HCl is used as digestion buffer, followed by vacuum evaporation at elevated temperatures, or samples are stored under alkaline conditions. Following the practical considerations of this paper can contribute to increasing the reliability of future analytical measurements.

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利用 HPLC-MS 对硫酸肝素二糖进行精确分析的样品处理步骤研究。

肝素硫酸盐（HSs）是一类特殊的糖胺聚糖（GAGs），是细胞信号的重要参与者。GAGs 的分析表征需要复杂的样品制备工作流程。虽然对硫酸软骨素类 GAG 进行了详细的稳定性和回收研究，但有关 HS 的文献在这方面并不完整。因此，我们的目的是系统地研究在使用高效液相色谱-质谱法进行二糖分析时可能影响 HS 样品稳定性和回收率的各种参数。首先，研究了真空蒸发和冷冻的影响。接着，分析了在温度、pH 值、消化缓冲液、进样溶剂和储存容器等多种条件下 HS 双糖的储存稳定性。我们确定了影响 HS 双糖稳定性和回收率的几个关键参数。我们得出结论，如果使用 Tris-HCl 作为消化缓冲液，然后在高温下进行真空蒸发，或将样品储存在碱性条件下，预计样品会大量流失。遵循本文的实际考虑有助于提高未来分析测量的可靠性。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

ELECTROPHORESIS 生物-分析化学

CiteScore

6.30

自引率

13.80%

发文量

244

审稿时长

1.9 months

期刊介绍： ELECTROPHORESIS is an international journal that publishes original manuscripts on all aspects of electrophoresis, and liquid phase separations (e.g., HPLC, micro- and nano-LC, UHPLC, micro- and nano-fluidics, liquid-phase micro-extractions, etc.). Topics include new or improved analytical and preparative methods, sample preparation, development of theory, and innovative applications of electrophoretic and liquid phase separations methods in the study of nucleic acids, proteins, carbohydrates natural products, pharmaceuticals, food analysis, environmental species and other compounds of importance to the life sciences. Papers in the areas of microfluidics and proteomics, which are not limited to electrophoresis-based methods, will also be accepted for publication. Contributions focused on hyphenated and omics techniques are also of interest. Proteomics is within the scope, if related to its fundamentals and new technical approaches. Proteomics applications are only considered in particular cases. Papers describing the application of standard electrophoretic methods will not be considered. Papers on nanoanalysis intended for publication in ELECTROPHORESIS should focus on one or more of the following topics: • Nanoscale electrokinetics and phenomena related to electric double layer and/or confinement in nano-sized geometry • Single cell and subcellular analysis • Nanosensors and ultrasensitive detection aspects (e.g., involving quantum dots, "nanoelectrodes" or nanospray MS) • Nanoscale/nanopore DNA sequencing (next generation sequencing) • Micro- and nanoscale sample preparation • Nanoparticles and cells analyses by dielectrophoresis • Separation-based analysis using nanoparticles, nanotubes and nanowires.