The BUME method: a new rapid and simple chloroform-free method for total lipid extraction of animal tissue

Abstract

In this study we present a simple and rapid method for tissue lipid extraction. Snap-frozen tissue (15–150 mg) is collected in 2 ml homogenization tubes. 500 μl BUME mixture (butanol:methanol [3:1]) is added and automated homogenization of up to 24 frozen samples at a time in less than 60 seconds is performed, followed by a 5-minute single-phase extraction. After the addition of 500 μl heptane:ethyl acetate (3:1) and 500 μl 1% acetic acid a 5-minute two-phase extraction is performed. Lipids are recovered from the upper phase by automated liquid handling using a standard 96-tip robot. A second two-phase extraction is performed using 500 μl heptane:ethyl acetate (3:1). Validation of the method showed that the extraction recoveries for the investigated lipids, which included sterols, glycerolipids, glycerophospholipids and sphingolipids were similar or better than for the Folch method. We also applied the method for lipid extraction of liver and heart and compared the lipid species profiles with profiles generated after Folch and MTBE extraction. We conclude that the BUME method is superior to the Folch method in terms of simplicity, through-put, automation, solvent consumption, economy, health and environment yet delivering lipid recoveries fully comparable to or better than the Folch method.

A 7‐day high‐fat, high‐calorie diet induces fibre‐specific increases in intramuscular triglyceride and perilipin protein expression in human skeletal muscle

“…Approximately 5 mg of freeze-dried muscle tissue (20-30 mg wet weight) was used for the lipid composition analysis using the butanol-methanol (3:1; BUME) method (Lofgren et al 2016). Briefly BUME solution was added to samples at −20°C and combined tissue homogenisation and lipid extraction was then performed using a Mixer Mill 301 instrument (Retsch GmbH, Haan, Germany).…”

“…Total lipid extracts were stored in chloroform-methanol (2:1) at −20°C until further analysis. Prior to mass spectrometric analysis, DAG was fractionated using straight-phase HPLC and evaporative light-scattering detection as previously described (Lofgren et al 2016). For mass spectrometric analysis, total lipid extracts, as well as the DAG fractions, were diluted with internal standard-containing chloroform-methanol (1:2) with 5 mM ammonium acetate.…”

Protein kinase MST3 modulates lipid homeostasis in hepatocytes and correlates with nonalcoholic steatohepatitis in humans

“…Lipids were also extracted using the BUME method (29) and quantified using MS. The cholesteryl esters, TAGs, and phospholipids were quantified using direct infusion on a QTRAP 5500 mass spectrometer (Sciex, Concord, ON, Canada) equipped with a robotic nanoflow ion source, the TriVersa NanoMate (Advion BioSciences, Ithaca, NJ, USA), and ceramides were quantified using ultra-high performance LC-MS (30).…”

One- vs two-phase extraction: re-evaluation of sample preparation procedures for untargeted lipidomics in plasma samples

“…Trying to achieve these objectives, while avoiding the inherent problems of the two-phase extraction methods, one-phase lipid extractions have recently been developed [2, 3, 15, 16]. One-phase extractions focus on an “all-in-one-tube” approach eliminating the need for phase separation by denaturing proteins that are later removed by centrifugation.…”