Improved Dried Blood Spot-Based Metabolomics Analysis by a Postcolumn Infused-Internal Standard Assisted Liquid Chromatography-Electrospray Ionization Mass Spectrometry Method

Divyabharathi Chepyala, Han Chun Kuo, Kang Yi Su, Hsiao Wei Liao, San Yuan Wang, Surendhar Reddy Chepyala, Lin Chau Chang, Ching Hua Kuo

Research output: Contribution to journalArticle

Abstract

Dried blood spots (DBSs) have gained increasing attention recently with their growing importance in precision medicine. DBS-based metabolomics analysis provides a powerful tool for investigating new biomarkers. Until now, very few studies have discussed measures for improving analytical accuracy with the consideration of the special characteristics of DBSs. The present study proposed a postcolumn infused-internal standard (PCI-IS) assisted strategy to improve data quality for DBS-based metabolomics studies. An efficient sample preparation protocol with 80% acetonitrile as the extraction solvent was first established to improve the metabolite recovery. The PCI-IS assisted liquid chromatography-electrospray ionization mass spectrometry (LC-ESI-MS) method was used to simultaneously estimate the blood volume and correct the signal change caused by ion source contamination and the matrix effect to evaluate the spot volume effect and hematocrit (Hct) variation effect on target metabolites. Phenylalanine-d8 was selected as the single PCI-IS to correct the matrix effect. For calibration of errors caused by the blood volume difference, 75% of the test metabolites showed good correlation (R2 ≥ 0.9) between the spot volume and the signal intensity after PCI-IS correction compared to less than 50% metabolites with good correlation before calibration. The spot volume was further calibrated by the same PCI-IS. Investigation of the Hct variation effect on target metabolites revealed that it affected the concentrations of metabolites in the DBS samples depending on their abundance in the red blood cell (RBC) or plasma; it is essential to preinvestigate the distribution of metabolites in blood to minimize the comparison bias in metabolomics studies. Finally, the PCI-IS assisted method was applied to study acetaminophen-induced liver toxicity. The results indicated that the proposed PCI-IS strategy could effectively remove analytical errors and improve the data quality, which would make the DBS-based metabolomics more feasible in real-world applications.

Original languageEnglish
JournalAnalytical Chemistry
DOIs
Publication statusPublished - Jan 1 2019

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Electrospray ionization
Liquid chromatography
Mass spectrometry
Blood
Metabolites
Metabolomics
Calibration
Biomarkers
Solvent extraction
Ion sources
Acetaminophen
Phenylalanine
Liver
Medicine
Toxicity
Contamination
Cells
Plasmas
Recovery

ASJC Scopus subject areas

  • Analytical Chemistry

Cite this

Improved Dried Blood Spot-Based Metabolomics Analysis by a Postcolumn Infused-Internal Standard Assisted Liquid Chromatography-Electrospray Ionization Mass Spectrometry Method. / Chepyala, Divyabharathi; Kuo, Han Chun; Su, Kang Yi; Liao, Hsiao Wei; Wang, San Yuan; Chepyala, Surendhar Reddy; Chang, Lin Chau; Kuo, Ching Hua.

In: Analytical Chemistry, 01.01.2019.

Research output: Contribution to journalArticle

Chepyala, Divyabharathi ; Kuo, Han Chun ; Su, Kang Yi ; Liao, Hsiao Wei ; Wang, San Yuan ; Chepyala, Surendhar Reddy ; Chang, Lin Chau ; Kuo, Ching Hua. / Improved Dried Blood Spot-Based Metabolomics Analysis by a Postcolumn Infused-Internal Standard Assisted Liquid Chromatography-Electrospray Ionization Mass Spectrometry Method. In: Analytical Chemistry. 2019.
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abstract = "Dried blood spots (DBSs) have gained increasing attention recently with their growing importance in precision medicine. DBS-based metabolomics analysis provides a powerful tool for investigating new biomarkers. Until now, very few studies have discussed measures for improving analytical accuracy with the consideration of the special characteristics of DBSs. The present study proposed a postcolumn infused-internal standard (PCI-IS) assisted strategy to improve data quality for DBS-based metabolomics studies. An efficient sample preparation protocol with 80{\%} acetonitrile as the extraction solvent was first established to improve the metabolite recovery. The PCI-IS assisted liquid chromatography-electrospray ionization mass spectrometry (LC-ESI-MS) method was used to simultaneously estimate the blood volume and correct the signal change caused by ion source contamination and the matrix effect to evaluate the spot volume effect and hematocrit (Hct) variation effect on target metabolites. Phenylalanine-d8 was selected as the single PCI-IS to correct the matrix effect. For calibration of errors caused by the blood volume difference, 75{\%} of the test metabolites showed good correlation (R2 ≥ 0.9) between the spot volume and the signal intensity after PCI-IS correction compared to less than 50{\%} metabolites with good correlation before calibration. The spot volume was further calibrated by the same PCI-IS. Investigation of the Hct variation effect on target metabolites revealed that it affected the concentrations of metabolites in the DBS samples depending on their abundance in the red blood cell (RBC) or plasma; it is essential to preinvestigate the distribution of metabolites in blood to minimize the comparison bias in metabolomics studies. Finally, the PCI-IS assisted method was applied to study acetaminophen-induced liver toxicity. The results indicated that the proposed PCI-IS strategy could effectively remove analytical errors and improve the data quality, which would make the DBS-based metabolomics more feasible in real-world applications.",
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