Features of Sample Preparation of Cell Culture Samples for Metabolomic Screening by LC-MS/MS | Sciact - CRIS-system of Boreskov Institute of Catalysis

Features of Sample Preparation of Cell Culture Samples for Metabolomic Screening by LC-MS/MS Full article

Journal

Journal of Pharmaceutical and Biomedical Analysis
ISSN: 2095-1779

Output data

Year: 2026, Volume: 267, Article number : 117146, Pages count : 12 DOI: 10.1016/j.jpba.2025.117146

Tags

Cell culture metabolomics; HPLC-MS/MS standardization; Cell number optimization; Targeted metabolomics; HILIC; RP LC

Authors

Basov Nikita V. ^1,2,3 , Butikova Ekaterina A. ⁴ , Sotnikova Maria A. ^1,2,3,4 , Razumov Ivan A. ⁵ , Sotnikova Yulia S. ^1,3,6 , Patrushev Yuriy V. ^3,6 , Rogachev Artem D. ^1,2 , Salakhutdinov Nariman F. ¹ , Pokrovsky Andrey G. ²

Affiliations

1	N.N. Vorozhtsov Novosibirsk Institute of Organic Chemistry, Siberian Branch of the Russian Academy of Sciences, Novosibirsk, Russia
2	Novosibirsk State University. Institute of Medicine and Medical Technologies, Novosibirsk, Russia
3	Novosibirsk State University. Department of Natural Sciences, Novosibirsk, Russia
4	Research Institute of Clinical and Experimental Lymphology, Branch of the Institute of Cytology and Genetics, Siberian Branch of the Russian Academy of Sciences, Novosibirsk, Russia
5	Institute of Cytology and Genetics, Siberian Branch of the Russian Academy of Sciences, Novosibirsk, Russia
6	Boreskov Institute of Catalysis, Siberian Branch of the Russian Academy of Sciences, Novosibirsk, Russia

Funding (5)

1	Ministry of Science and Higher Education of the Russian Federation	FWNR-2022-0023
2	Ministry of Science and Higher Education of the Russian Federation	FWUR-2024-0032
3	Ministry of Science and Higher Education of the Russian Federation	075-00365-25-00
4	Ministry of Science and Higher Education of the Russian Federation	FSUS-2025-0012
5	Ministry of Science and Higher Education of the Russian Federation	FWNR-2025-0016 (125031103454-7)(075-03-2025-636)

Metabolomic analysis has become an essential tool in the life sciences, providing insights into cellular metabolism. However, preparing cell cultures for metabolomic screening remains challenging, especially with samples containing variable cell numbers. Standardized and reproducible protocols are required to ensure reliable data while maintaining compatibility with high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). Using melanoma cell lines SK-MEL-28 (human) and B16 (mouse) as models, we developed and optimized a convenient sample preparation protocol for metabolomic screening by HPLC-MS/MS. The study is focused on optimizing key steps, including cell lysis, metabolite extraction, and normalization strategies for accurate semiquantitative analysis. The effects of cell count on metabolomic coverage and detection sensitivity were evaluated using hydrophilic interaction liquid chromatography (HILIC) and reversed-phase (RP) chromatography. The protocol enables efficient detection of several metabolite classes from samples containing as few as 10,000 cells. The optimal cell count for reliable analysis was found to be 400,000 – 500,000 cells, ensuring consistent and reproducible detection within the method’s analytical coverage. Our findings emphasize the importance of cell size and number in metabolomic studies, as larger cells provide improved metabolomic coverage. Moreover, metabolites exhibited varying detection limits, highlighting the need to adjust sample preparation strategies according to metabolite characteristics. The proposed protocol offers a robust and reproducible approach for the metabolomic screening of adherent melanoma cell cultures by HPLC-MS/MS and can be adapted for non-adherent and other cell types. Balancing sensitivity, reproducibility, and feasibility, this method provides a standardized solution for cell metabolomic studies in pharmacometabolomics, cancer research, and related fields.

Basov N.V. , Butikova E.A. , Sotnikova M.A. , Razumov I.A. , Sotnikova Y.S. , Patrushev Y.V. , Rogachev A.D. , Salakhutdinov N.F. , Pokrovsky A.G.
Features of Sample Preparation of Cell Culture Samples for Metabolomic Screening by LC-MS/MS
Journal of Pharmaceutical and Biomedical Analysis. 2026. V.267. 117146 :1-12. DOI: 10.1016/j.jpba.2025.117146 WOS Scopus РИНЦ AN PMID OpenAlex

Submitted:	Jun 11, 2025
Accepted:	Sep 13, 2025
Published online:	Sep 15, 2025
Published print:	Jan 1, 2026

Web of science:	WOS:001584074500002
Scopus:	2-s2.0-105016313330
Elibrary:	82893525
Chemical Abstracts:	2025:2312177
PMID:	40972480
OpenAlex:	W4414230274

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