Quantification of Diacylglycerol by Mass Spectrometry

Abstract

Diacylglycerol (DAG) is an important intermediate of lipid metabolism and a component of phospholipase C signal transduction. Quantification of DAG in plant membranes represents a challenging task because of its low abundance. DAG can be measured by direct infusion mass spectrometry (MS) on a quadrupole time-of-flight mass spectrometer after purification from the crude plant lipid extract via solid-phase extraction on silica columns. Different internal standards are employed to compensate for the dependence of the MS and MS/MS signals on the chain length and the presence of double bonds in the acyl moieties. Thus, using a combination of single MS and MS/MS experiments, quantitative results for the different molecular species of DAGs from Arabidopsis can be obtained.

Tracking synthesis and turnover of triacylglycerol in leaves

“…First, ~77% of DAG molecular species in Arabidopsis leaves do not have two double bonds. The predominant species with two double bonds is SD (16:0/18:2) and represents only ~23% ( Vom Dorp et al ., 2013 ). Although these data do not distinguish the distribution of DAG between plastid and extraplastidial membranes, the low level of 16:0/18:1 molecular species and almost complete absence of 16:3 species implies that most leaf DAG is extraplastidial.…”

The Lotus japonicus acyl‐acyl carrier protein thioesterase FatM is required for mycorrhiza formation and lipid accumulation of Rhizophagus irregularis

“…Thirty‐five days after inoculation, the medium was dissolved with 10 m m sodium citrate and the ERM and the roots harvested, washed in sterile water and used for lipid extraction. To extract MAGs, total lipid extracts from colonized D. carota roots, R. irregularis ERM and colonized or mock‐treated Lotus roots were dried, dissolved in hexane and the MAGs purified by solid‐phase extraction on silica columns (vom Dorp et al ., ). The MAGs were eluted with diethylether then dried by evaporation.…”

“…After 12 h, 2 ml of the culture containing the cells was removed and boiled at 100°C for 10 min. Lipids were extracted from the culture (including cells and medium) with n ‐hexane and the hexane phase was used for purification of free fatty acids by solid‐phase extraction (vom Dorp et al ., ). Free fatty acids were converted into methyl esters with 1 m methanolic HCl and quantified via GC using pentadecanoic acid (15:0) as an internal standard (Browse et al ., ).…”

Effects of repeated drought stress on the physiological characteristics and lipid metabolism of Bombax ceiba L. during subsequent drought and heat stresses

“…However, this should be further confirmed through determination of the metabolic reprogramming of carbohydrates and the related enzyme activities in those processes. DAG and PA are the intermediates of lipid biosynthesis and also important second messengers that help to regulate cell functions [43,51,52]. Without degradation of other lipid classes, the substantial production of DAG under D3 treatment (Additional file 3) may have been due to an increased de novo synthesis.…”