Plasma Lipidomic Profiling Method Based on Ultrasound Extraction and Liquid Chromatography Mass Spectrometry

Abstract

Lipidomics is an emerging field in biomedical research that includes the analysis of all the lipids present in complex biological samples. To evaluate the chemical and biological diversity of lipids, lipid extraction is usually the first step toward lipidomics analysis. Nevertheless, sample preparation is still a time-consuming and error prone analytical step. Therefore, the development of simple and robust methods suitable for high-throughput lipid analysis is of great interest. This study presents a new method for exhaustive lipid fingerprinting of human blood plasma samples based on the employment of methyl tert-butyl ether (MTBE) and ultrasound (US) energy combined with liquid chromatography-electrospray ionization quadrupole-time-of-flight mass spectrometry (LC-ESIqToF-MS). First, the MTBE-US extraction step was optimized by means of experimental design methodology. After the optimization step, a comparative study was performed to assess the suitability of the proposed method. The new method allowed extraction time to be reduced to half, in comparison with previously reported methods. The proposed method also allowed increasing extraction repeatability (with RSDs below 5.55%) and efficiency (recoveries higher than 70% were obtained for all lipids evaluated). Moreover, the new proposed method enables more than 800 different features to be detected. Thus, the overall number of lipids identified with the databases for this novel extraction method (352) was the highest of the evaluated methods. The efficiency, precision, and feature detection capacity of the proposed method confirmed its suitability for the evaluation of the lipid profile of human blood plasma samples. Moreover, taking into account its simplicity, low time consumption, and compatibility with automation, the new proposed method could be a suitable alternative to previously reported methods for use in laboratories for comprehensive lipidomic profiling.

Lipidomics Reveals a Tissue-Specific Fingerprint

“…Samples were immersed in a water bath (ATU Ultrasonidos, Valencia, Spain) with an ultrasound frequency and power of 40 kHz and 100 W, respectively, at 10°C for 30 min. Then, 25 μL of miliQ water were added to the mixture, and organic phase was separated by centrifugation (1,400 g) at 10°C for 10 min (Pizarro et al, 2013). Lipid extracts, contained in the upper phase, were collected and subjected to mass spectrometry.…”

Free Fatty Acids Profiles Are Related to Gut Microbiota Signatures and Short-Chain Fatty Acids

“…Free fatty acids were analyzed after a methyl- tert -butylether (MTBE)-based extraction protocol as previously described (24) with slight modifications. Briefly, 100 µl serum samples were spiked with 5 µl of internal standard (600 ppm heptadecanoic acid).…”

Recent advances in lipidomics for disease research

“…developed and validated a new one‐phase extraction method, demonstrating that the mixture of methanol/chloroform/methyl tert ‐butyl ether (MTBE) provided a recovery very close to 100% for all nine lipid classes in human serum. Besides, Pizarro et al . also used MTBE as extraction solvent combined with ultrasound‐assisted extraction to present a fast and precise method for exhaustive fingerprinting lipidomics.…”