- A major challenge of lipidomics is to determine and quantify the precise content of complex lipidomes to the exact lipid molecular species. Often, multiple methods are needed to achieve sufficient lipidomic coverage to make these determinations. Multiplexed targeted assays offer a practical alternative to enable quantitative lipidomics amenable to quality control standards within a scalable platform. Herein, we developed a multiplexed normal phase liquid chromatography-hydrophilic interaction chromatography multiple reaction monitoring method that quantifies lipid molecular species across over 20 lipid classes spanning wide polarities in a single 20-min run.
- Phospholipids with a choline head group are an abundant component of cellular membranes and are involved in many important biological functions. For studies on the cell biology and metabolism of these lipids, traceable analogues where propargylcholine replaces the choline head group have proven useful. We present a novel method to analyze propargylcholine phospholipids by MS. The routine employs 1-radyl-2-lyso-sn-glycero-3-phosphopropargylcholines as labeled lysophosphatidylcholine precursors, which upon cellular conversion direct the traceable tag with superb specificity and efficiency to the primary target lipid class.
- Lysophosphatidic acid (LPA) is a potent signaling lipid, and state-dependent alterations in plasma LPA make it a promising diagnostic marker for various diseases. However, plasma LPA concentrations vary widely among reports, even under normal conditions. These variations can be attributed, at least in part, to the artificial metabolism of LPA after blood collection. Here, we aimed to develop an optimized plasma preparation method that reflects the concentration of LPA in the circulating blood. The main features of the devised method were suppression of both LPA production and degradation after blood collection by keeping whole blood samples at low temperature followed by the addition of an autotaxin inhibitor to plasma samples.
- Lipidomics data require consideration of ions with near-identical masses, which comprises among others the Type-II isotopic overlap. This overlap occurs in series of lipid species differing only by number of double bonds (DBs) mainly because of the natural abundance of 13C-atoms. High-resolution mass spectrometry, such as Fourier-transform mass spectrometry (FTMS), is capable of resolving Type-II overlap depending on mass resolving power. In this work, we evaluated FTMS quantification accuracy of lipid species affected by Type-II overlap.
- Lipases constitute an important class of water-soluble enzymes that catalyze the hydrolysis of hydrophobic triacylglycerol (TAG). Their enzymatic activity is typically measured using multistep procedures involving isolation and quantification of the hydrolyzed products. We report here a new fluorescence method to measure lipase activity in real time that does not require the separation of substrates from products. We developed this method using adipose triglyceride lipase (ATGL) and lipoprotein lipase (LpL) as model lipases.