Chemical Labeling Assisted Detection and Identification of Short Chain Fatty Acid Esters of Hydroxy Fatty Acid in Rat Colon and Cecum Contents

Abstract

Branched fatty acid esters of hydroxy fatty acids (FAHFAs) are novel endogenous lipids with important physiological functions in mammals. We previously identified a new type of FAHFAs, named short-chain fatty acid esterified hydroxy fatty acids (SFAHFAs), with acetyl or propyl esters of hydroxy fatty acids of carbon chains, C ≥ 20. However, sensitive determination of SFAHFAs is still a challenge, due to their high structural similarity and low abundance in biological samples. This study employs one-step chemical derivatization following total lipid extraction using 2-dimethylaminoethylamine (DMED) for enhanced detection of SFAHFAs. The labeled extracts were subjected to ultrahigh performance liquid chromatography coupled to linear ion trap quadrupole-Orbitrap mass spectrometry (UHPLC/LTQ-Orbitrap MS). Our results demonstrated that the detection sensitivities of SFAHFAs increased after DMED labeling, and is highly helpful in discovering six additional novel SFAHFAs in the cecum and colon contents of WKAH/HKmSlc rats fed with normal and high-fat diet (HFD). The identified DMED labeled SFAHFAs were characterized by their detailed MS/MS analysis, and their plausible fragmentation patterns were proposed. The concentrations of SFAHFAs were significantly reduced in the cecum of HFD group compared to the control. Hence, the proposed method could be a promising tool to apply for the enhanced detection of SFAHFAs in various biological matrices, which in turn facilitate the understanding of their sources, and physiological functions of these novel lipids.

Polyunsaturated fatty acids and fatty acid-derived lipid mediators: Recent advances in the understanding of their biosynthesis, structures, and functions

“…Branched FAHFAs are endogenously produced in insects [ 250 ], and mammals, such as rodents [ 248 , 251 ], caribou and moose [ 252 ], and humans [ 251 , 253 ]. In humans, white adipose tissue (WAT) represents the major site of FAHFA synthesis [ 248 , 253 – 255 ], although they are also found in blood [ 256 , 257 ] and other tissues, such as the liver [ 248 , 258 ], kidneys [ 248 , 258 , 259 ], large intestine [ 260 ], pancreas [ 248 ], lungs [ 258 ], thymus [ 258 ], and heart [ 258 ], albeit to a lesser extent. Branched FAHFAs are also naturally occurring substances found in microalgae [ 261 ], breast milk [ 262 ] and foods, such as cereals, fruits, vegetables, oils, eggs and meat [ 251 , 263 – 265 ].…”

Detection and Structural Characterization of SFAHFA Homologous Series in Mouse Colon Contents by LTQ-Orbitrap-MS and Their Implication in Influenza Virus Infection

“…The lipids were extracted from the colon contents by the method established earlier in our laboratory with minor modifications. , In brief, about 20–100 mg of colon contents was weighed into a 1.5 mL Eppendorf tube, and approximately 5–6 ceramic beads (1.4 mm, catalog no. 15-340-159, Fisherbrand) and 10 vol of cold methanol were added.…”

“…21−23 The acetate-and propionate-derived SFAHFAs identified in our previous study were also characterized by high-resolution mass spectrometry and improved detection sensitivity by chemical labeling. 14,24 The identified SFAHFAs were decreased significantly in colon contents of rats fed a high-fat diet, 14 demonstrating their biological importance in the gut. However, so far, only a limited number of SFAHFAs with HFA ≥C20 were reported, and their double-bond position was not characterized.…”

“…Although 2-HFAs are major fatty acyl components of mammalian sphingolipids, identification of 2-HFA-derived FAHFAs are limited in the literature. Analysis and characterization of FAHFAs are extensively performed by liquid chromatography–mass spectrometry (LC/MS). ,, Further, chemical derivatization enhanced detection techniques were coupled to LC/MS. − The acetate- and propionate-derived SFAHFAs identified in our previous study were also characterized by high-resolution mass spectrometry and improved detection sensitivity by chemical labeling. , The identified SFAHFAs were decreased significantly in colon contents of rats fed a high-fat diet, demonstrating their biological importance in the gut. However, so far, only a limited number of SFAHFAs with HFA ≥C20 were reported, and their double-bond position was not characterized.…”

Potential favourable health effects of some dietary uncommon fatty acids

“…In lung, kidney, thymus, liver and heart FAHFAs content is rather in pg/g (Zhu et al, 2017). Short-chain FAHFAs are in a concentration range from 0.84 to 57 pmoles/mg in the large intestine (Gowda et al, 2020b(Gowda et al, , 2020c.…”