Development of an analytical method for human blood triglycerides using triple quadrupole mass spectrometer

Significance of the Topic

Triglycerides in human blood serve as key energy carriers and indicators of metabolic health. Accurate profiling of individual triglyceride species can reveal early signs of cardiovascular and metabolic disorders. Conventional clinical assays quantify only total triglyceride levels, overlooking the molecular diversity that may hold diagnostic value.

Objectives and Study Overview

This work aims to develop a rapid, sensitive LC-MS/MS method for comprehensive quantitation of 47 distinct triglycerides in human plasma and serum. The study demonstrates a high-throughput workflow capable of processing 130 samples per day and applies principal component analysis (PCA) to distinguish subtle lipidomic differences among biological samples.

Used Instrumentation

• UHPLC: Shimadzu Nexera X2 system with Shim-pack Velox C18 column (50 × 2.1 mm, 2.7 µm) at 45 °C
• Mass Spectrometer: Shimadzu LCMS-8060 triple quadrupole
• Ionization: ESI in positive mode; MRM transitions for neutral loss scanning

Methodology and Sample Preparation

Plasma or serum (20 µL) was spiked with glyceryl trilinolenate internal standard and extracted with methanol/butanol (1:1). After centrifugation, the supernatant was diluted and injected (3 µL) into the LCMS-8060. Chromatographic separation used a gradient of 20 mM ammonium formate (water) and 2-propanol/acetonitrile, achieving elution of all targets within 8 minutes.

Main Results and Discussion

MRM chromatograms clearly resolved 47 triglyceride species; TG 52:2 exhibited the highest response in plasma. Retention times decreased with increasing double bond count. Data normalization to the internal standard enabled robust quantitation. PCA on 195 MRM features separated two plasma samples and one serum sample into distinct clusters (p < 0.01 for 109 lipid components), highlighting sample-specific lipid signatures.

Benefits and Practical Applications

This LC-MS/MS approach permits detailed triglyceride profiling with high throughput, offering enhanced insight into fatty acid composition versus standard total TG assays. It supports biomarker discovery for metabolic and cardiovascular disease research, quality control in blood lipid laboratories, and nutritional studies monitoring dietary fatty acid incorporation.

Future Trends and Potential Applications

Advances may include automation of sample handling, expansion to cover oxidized and odd-chain triglycerides, and integration with data-driven lipidomic platforms. Coupling this method with clinical cohorts could refine personalized risk assessment models and foster new therapeutic targets.

Conclusion

An LC-MS/MS method using a triple quadrupole MS and UHPLC separation was established for rapid, high-throughput analysis of 47 human blood triglycerides. Its sensitivity and speed position it as a valuable tool for clinical lipidomics and biomarker research.

References

• Liebisch G, Fahy E, Aoki J, et al. Update on LIPID MAPS classification, nomenclature, and shorthand notation for MS-derived lipid structures. J Lipid Res. 2020;61(12):1539-1555. doi:10.1194/jlr.S120001025