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Lipid species patterns are conserved within cells to maintain physicochemical properties of membranes and cellular functions. We present the lipidome, including sterols, glycerolipids (GLs), glycerophospholipids (GPLs), and sphingolipids (SLs), of primary ex vivo differentiated (I) white, (II) brite, and (III) brown adipocytes derived from primary preadipocytes isolated from (I) epididymal white, (II) inguinal white, and (III) intrascapular brown adipose tissue. Quantitative lipidomics revealed significantly decreased fractions of phosphatidylcholine (PC) and phosphatidylethanolamine (PE), with longer (C > 36) and more polyunsaturated species, as well as lower levels of cardiolipin (CL) in white than in brite and brown adipocytes. Together, the brite and brown lipidome was comparable and indicates differences in membrane lipid packing density compared with white adipocytes. Changes in ceramide species profile could be related to the degree of browning. Beta-adrenergic stimulation of brown adipocytes led to generation of saturated lyso-PC (LPC) increasing uncoupling protein (UCP) 1-mediated leak respiration. Application of stable isotope labeling showed that LPC formation was balanced by an increased de novo synthesis of PC.

Lipids are a ubiquitous class of structurally complex molecules involved in various biological processes. In the fast-growing field of lipidomics, preanalytical issues are frequently neglected. Here, we investigated the stability of lipid profiles of murine liver, brain, lung, heart, and spleen homogenates by quantitative flow injection analysis using tandem mass spectrometry and high-resolution mass spectrometry. Storage of tissue homogenates at room temperature showed substantial alterations of the lipid profiles reflecting lipolytic action. Therefore, ratios of ceramide to sphingomyelin, lysophosphatidylethanolamine to phosphatidylethanolamine, lysophosphatidylcholine to phosphatidylcholine, and diglyceride to triglyceride were applied to monitor sample stability and the effect of sodium dodecyl sulfate (SDS) as a potential stabilizing agent. The addition of SDS led to a concentration-dependent stabilization of lipid profiles in liver, brain, and heart homogenates, while in lung and spleen homogenates, in particular, the lysophosphatidylethanolamine to phosphatidylethanolamine ratio increased upon addition of SDS. In conclusion, we demonstrated that lipid class ratios reflecting lipolytic activity could be applied to evaluate both the stability of samples and the influence of stabilizers.