Simple, Rapid Lipidomic Analysis of Triacylglycerols in Bovine Milk by Infusion‐Electrospray Mass Spectrometry

Abstract

Bovine milk is a complex mixture of lipids, proteins, carbohydrates, and other factors of which lipids comprise 3–5% of the total mass. Rapid analysis and characterization of the triacylglycerols (TAG) that comprise about 95% of the total lipid is daunting given the numerous TAG species. In the attached methods paper, we demonstrate an improved method for identifying and quantifying TAG species by infusion‐based “shotgun” lipidomics. Because of the broad range of TAG species in milk, a single internal standard was insufficient for the analysis and required sectioning the spectrum into three portions based upon mass range to provide accurate quantitation of TAG species. Isobaric phospholipid interferences were removed using a simple dispersive solid‐phase extraction step. Using this method, > 100 TAG species were quantitated by acyl carbon number and desaturation level in a sample of commercially purchased bovine milk.

Lipidomic Analysis of TRPC1 Ca2+-Permeable Channel-Knock Out Mouse Demonstrates a Vital Role in Placental Tissue Sphingolipid and Triacylglycerol Homeostasis Under Maternal High-Fat Diet

“…Data were collected on an AB Sciex 5500 QTRAP hybrid mass spectrometer equipped with a Turbo V electrospray ion source and SelexION ion mobility device (AB Sciex, Framingham, MA, USA). Samples were infused using a Shimadzu Prominence UPLC system (Shimadzu Scientific Instruments, Columbia, MD, USA) equipped with an LC20XR autosampler (50 µL stainless steel sample loop) and two solvent delivery units following a configuration modified from Bukowski and Picklo as detailed in the Supplemental Information ( 19 ).…”

“…Multiple instrument modes were used based upon lipid class and preparation ( Table S2 ). TAG were characterized and quantitated by brutto structure as ammoniated cations, [TAG + NH 4 ] + in enhanced mass spectrum (EMS) mode as previously published ( 19 , 20 ). Cholesterol esters (CE) were assayed by neutral loss scan for 20 fatty acid neutral losses with confirmatory detection of the cholesterol head group by product ion scan for mass-to-charge ratio (m/z) = 369, as previously published ( Table S3 ) ( 20 , 21 ).…”

“…Mass spectra for all lipid class were examined manually to confirm brutto structure assignment. Isotopic and ionization correction factors were determined as previously described and quantitation of target species was performed using LipidView software (AB Sciex, Framingham, MA, USA) ( 19 , 20 , 22 , 25 ). Values were normalized to tissue wet weight.…”

Quantifying Sphingomyelin in Dairy through Infusion-Based Shotgun Mass Spectrometry with Lithium-Ion-Induced Fragmentation

“…As a result of differential ionization of SM species as functions of the acyl carbon number ( C ) and desaturation level ( N ), an equimolar calibration solution was used to calibrate the ESI source. , Preparation of similar ESI calibration solutions for PC is documented elsewhere . Stock solutions of seven SM species with a d18:1 sphingosine base and acyl fatty acids ranging from C14 to C24 were prepared in 1:1 methanol/chloroform (Table S1 of the Supporting Information).…”

Ammonia Concentration in the Eluent Influences Fragmentation Pattern of Triacylglycerols in Mass Spectrometry Analysis

“…Additionally, methodologies encompassing shot-gun or direct infusion versus separation by liquid chromatography before MS detection are currently used and are a matter of debate regarding their efficiency [ 5 , 6 , 16 ]. Therefore, active research concerning the influence of other factors on the fragmentation information quality, as well as the application of diverse methodologies, is ongoing at present regarding TAG analysis [ 17 , 18 , 19 , 20 , 21 , 22 , 23 , 24 , 25 ].…”