Comparing nitric acid treatment and microwave digestion for efficiency of metal extraction from bioprocess samples

IF 2.6 4区生物学 Q2 BIOCHEMICAL RESEARCH METHODS

Analytical biochemistry Pub Date : 2024-05-03 DOI:10.1016/j.ab.2024.115555

Adil Mohammad, Chikkathur N. Madhavarao

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引用次数: 0

Abstract

Metal ions may act as enzyme cofactors and influence the kinetics of biochemical reactions that may also influence the biological production of therapeutic proteins and quality attributes such as glycosylation. Because sample preparation is a significant step in the reliable analysis of metals, we compared two sample preparation procedures for metal analysis of bioreactor culture media samples by ICP-MS: (i) samples were diluted in 2 % nitric acid (treatment with nitric acid, TNA); and (ii) samples were mixed with equal volume of 5 % nitric acid and closed vessel digestion was performed in a microwave (closed vessel digestion, CVD). In the comparison of extraction efficiencies between TNA and CVD procedures, CVD showed better extraction for Ca and Cu among bulk metals (∼30 %) and for Ni among the trace metals (∼65 %) for the bioreactor broth supernatant samples. For the cell pellet samples, the CVD procedure was found to be better for extraction of Fe (∼65 % more) among bulk metals, Zn (∼20 % more) among minor metals and Co (∼60 % more) and Ni (∼45 % more) among trace metals. Differences between the two procedures were less than 10 % and TNA was better for all other metals quantified from both supernatant samples and cell pellet samples. The current study helps bring more clarity to the methodology on comprehensive metal analysis to monitor and maintain trace metal content for biologics production.

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比较硝酸处理和微波消解从生物处理样品中提取金属的效率。

金属离子可作为酶的辅助因子，影响生化反应的动力学，也可能影响治疗蛋白质的生物生产和质量属性（如糖基化）。由于样品制备是可靠分析金属的重要步骤，我们比较了用 ICP-MS 对生物反应器培养基样品进行金属分析的两种样品制备程序：(i) 用 2% 硝酸稀释样品（硝酸处理，TNA）；(ii) 将样品与等体积的 5% 硝酸混合，然后在微波中进行密闭容器消化（密闭容器消化，CVD）。在比较 TNA 和 CVD 两种萃取方法的萃取效率时，在生物反应器肉汤上清液样品中，CVD 对大宗金属中的 Ca 和 Cu 的萃取效率更高（∼30%），对痕量金属中的 Ni 的萃取效率更高（∼65%）。在细胞团样品中，CVD 程序对大宗金属中铁的萃取效果更好（高出 65%），对小金属中锌的萃取效果更好（高出 20%），对微量金属中钴（高出 60%）和镍（高出 45%）的萃取效果更好。两种方法之间的差异小于 10%，而且 TNA 对上清液样品和细胞团样品中所有其他金属的定量效果更好。本研究有助于进一步明确全面金属分析的方法，以监测和维护生物制品生产中的痕量金属含量。

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

Analytical biochemistry 生物-分析化学

CiteScore

5.70

自引率

0.00%

发文量

283

审稿时长

44 days

期刊介绍： The journal''s title Analytical Biochemistry: Methods in the Biological Sciences declares its broad scope: methods for the basic biological sciences that include biochemistry, molecular genetics, cell biology, proteomics, immunology, bioinformatics and wherever the frontiers of research take the field. The emphasis is on methods from the strictly analytical to the more preparative that would include novel approaches to protein purification as well as improvements in cell and organ culture. The actual techniques are equally inclusive ranging from aptamers to zymology. The journal has been particularly active in: -Analytical techniques for biological molecules- Aptamer selection and utilization- Biosensors- Chromatography- Cloning, sequencing and mutagenesis- Electrochemical methods- Electrophoresis- Enzyme characterization methods- Immunological approaches- Mass spectrometry of proteins and nucleic acids- Metabolomics- Nano level techniques- Optical spectroscopy in all its forms. The journal is reluctant to include most drug and strictly clinical studies as there are more suitable publication platforms for these types of papers.