H OH metabolites OH Review Fatty Acyl Esters of Hydroxy Fatty Acid (FAHFA) LipidReview Families Fatty Acyl Esters of Hydroxy Fatty Acid (FAHFA) Paul L. Wood MetabolomicsLipid Unit,Families College of Veterinary Medicine, Lincoln Memorial University, 6965 Cumberland Gap Pkwy, Harrogate, TN 37752, USA; [email protected] Paul L. Wood Received: 15 November 2020; Accepted: 16 December 2020; Published: 17 December 2020 Metabolomics Unit, College of Veterinary Medicine, Lincoln Memorial University, 6965 Cumberland Gap Pkwy., Harrogate, TN 37752, USA; [email protected] Abstract:Received:Fatty 15 November Acyl esters 2020; of Accepted: Hydroxy 16 Fatty December Acids 2020; (FAHFA) Published: encompass 17 December three 2020 different lipid families which have incorrectly been classified as wax esters. These families include (i) Branched-chain FAHFAs,Abstract: involved Fatty Acyl in the esters regulation of Hydroxy of glucose Fatty Acids metabolism (FAHFA) and encompass inflammation, three different with acylation lipid of an internalfamilies branched-chainwhich have incorrectly hydroxy-palmitic been classified oras -stearicwax esters. acid; These (ii) families!-FAHFAs, include which (i) Branched- function as biosurfactantschain FAHFAs, in a involved number in ofthe biofluids, regulation areof gluc formedose metabolism via acylation and inflammation, of the !-hydroxyl with acylation group of very-long-chainof an internal fattybranched-chain acids (these hydroxy-palmitic lipids have also or -stearic been designated acid; (ii) ω-FAHFAs, as o-acyl which hydroxy function fatty as acids; biosurfactants in a number of biofluids, are formed via acylation of the ω-hydroxyl group of very- OAHFA); and (iii) Ornithine-FAHFAs are bacterial lipids formed by the acylation of short-chain long-chain fatty acids (these lipids have also been designated as o-acyl hydroxy fatty acids; 3-hydroxyOAHFA); fatty and acids (iii) andOrnithine-FAHFAs the addition of are ornithine bacterial to lipids the free formed carboxy by the group acylation of the of hydroxyshort-chain fatty 3- acid. The dihydroxyfferences fatty in acids biosynthetic and the addition pathways of ornithine and cellular to the functions free carboxy of these group lipid of the families hydroxy will fatty be acid. reviewed and comparedThe differences to wax in esters,biosynthetic which pathways are formed and by cell theular acylation functions of aof fatty these alcohol, lipid families not a hydroxy will be fatty acid.reviewed In summary, and compared FAHFA lipid to wax families esters, arewhich both are unique formed and by the complex acylation in theirof a fatty biosynthesis alcohol, not and a their biologicalhydroxy actions. fatty acid. We have In summary, only evaluated FAHFA the lipid tip families of the iceberg are both and unique much and more complex exciting in researchtheir is requiredbiosynthesis to understand and their these biological lipids actions. in health We and have disease. only evaluated the tip of the iceberg and much more exciting research is required to understand these lipids in health and disease. Keywords: FAHFA; OAHFA; ornithine; wax esters; !-hydroxylation Keywords: FAHFA; OAHFA; ornithine; wax esters; ω-hydroxylation 1. Introduction 1. Introduction In thisIn this review, review, we we present present an an abbreviated abbreviated overviewoverview of of three three major major Fatty Fatty Acyl Acyl esters esters of Hydroxy of Hydroxy FattyFatty Acids Acids (FAHFA) (FAHFA) lipid lipid families families which which individuallyindividually serve serve unique unique metabolic metabolic and/or and structural/or structural functionsfunctions (Figure (Figure1). Branched-chain 1). Branched-chain FAHFAs FAHFAs are essential are essential modulators modulators of metabolic of metabolic and inflammatory and healthinflammatory status. These health FAHFAs status. are These formed FAHFAs by the are acylation formed by of the medium acylation chain of medium (16 to 18 chain carbons) (16 to hydroxy 18 fattycarbons) acids. hydroxy fatty acids. (A) Branched-chain FAHFA 16:0/12-O-18:0 (LMFA07090007; C34H66O4; 538.4961). (B) ω-FAHFA 18:1/ω-O-32:1 (LMFA07090154; C50H94O4; 758.7152) Metabolites 2020, 10, x; doi: FOR PEER REVIEW Figure 1. Cont. www.mdpi.com/journal/metabolites Metabolites 2020, 10, 512; doi:10.3390/metabo10120512 www.mdpi.com/journal/metabolites Metabolites 2020, 10, 512 2 of 8 Metabolites 2020, 10, x FOR PEER REVIEW 2 of 9 (C) ORN-FAHFA 18:0/3-O-18:0 (LMFA08020226; C41H80N2O5; 680.6067) (D) WE 18:0/16:0 (LMFA07010049; C34H68O2; 508.5219) FigureFigure 1. Structures 1. Structures of fattyof fatty acyl acyl esters esters of of hydroxyhydroxy fa fattytty acids acids (FAHFA) (FAHFA) and and wax wax esters esters (WE). (WE). The acyl The acyl linkages are formed by the following reaction: R-OH + R′-COOH = R-O-CO-R′ +H2O. In eukaryotes, linkages are formed by the following reaction: R-OH + R0-COOH = R-O-CO-R0 +H2O. In eukaryotes, the majorthe major branched-chain branched-chain FAHFAs FAHFAs (A (A)) are are palmitic palmitic acidacid/hydroxystearic/hydroxystearic acid acid (16:0/O-18:0) (16:0/O-18:0) and andoleic oleic acid/hydroxystearicacid/hydroxystearic acid acid (18:1 (18:1/O-18:0),/O-18:0), with with thethe hydroxyl group group at atcarbons carbons 5, 9, 5, or 9, 12 or of 12 stearic of stearic acid. acid. The ω-FAHFAs, represented in (B), are unique in that the hydroxy fatty acids are long-chain fatty The !-FAHFAs, represented in (B), are unique in that the hydroxy fatty acids are long-chain fatty acids with a terminal (ω) hydroxyl group. In the case of ornithine (ORN)-FAHFA (C), the hydroxy acids with a terminal (!) hydroxyl group. In the case of ornithine (ORN)-FAHFA (C), the hydroxy fatty acid is consistently a 3-hydroxy fatty acid and the addition of ORN is to the free carboxyl group fattyof acid the is hydroxy consistently fatty acid a 3-hydroxy component fatty of th acide FAHFA. and the In additioncontrast, wax of ORN esters is (WE; to the D free) are carboxylformed by group of thethe hydroxy acylation fatty of a fatty acid alcohol, component resulting of the in the FAHFA. formation Incontrast, of a neutral wax lipid, esters which (WE; contrastsD) are with formed the by the acylationcharged FAHFA of a fatty lipids. alcohol, While resulting Lipid Maps in categorizes the formation FAHFAs of a as neutral wax esters, lipid, the which completely contrasts different with the chargedbiosynthetic FAHFA lipids.pathways While (i.e., Lipidfatty acid Maps vs. categorizes fatty alcohol FAHFAs addition) as indicates wax esters, that this the completelyis too simplified different biosyntheticand incorrect. pathways The Lipid (i.e., fattyMaps acidnumber vs. fattyand ex alcoholact mass addition) of the lipids indicates are also that provided. this is too simplified and incorrect. The Lipid Maps number and exact mass of the lipids are also provided. ω-FAHFAs are unique in that they are synthesized by the acylation of very-long-chain ω- !hydroxy-FAHFAs fatty are acids. unique This in thatsynthetic they are pathway synthesized requires by thecomplex acylation metabolic of very-long-chain machinery for! -hydroxythe fattybiosynthesis acids. This of synthetic very-long-chain pathway ω-hydroxy requires fatty complex acids (30 metabolic to 34 carbons). machinery ω-FAHFAs, for which the biosynthesis have also of very-long-chainbeen termed! o-acyl-hydroxy hydroxy fatty fatty acids acids (30 (OAHFA), to 34 carbons). are biosurfactants!-FAHFAs, in which the tear have film also of beenthe eye, termed amniotic fluid, semen, sperm, skin, and the vernix caseosa. o-acyl hydroxy fatty acids (OAHFA), are biosurfactants in the tear film of the eye, amniotic fluid, Ornithine-FAHFA are bacterial lipids synthesized by the acylation of short-chain 3-hydroxy semen,fatty sperm, acids skin,and the and addition the vernix of ornithine caseosa. to the free carboxy group of the hydroxy fatty acid. These Ornithine-FAHFAcomplex acylated ornithines are bacterial act as lipidsa permeability synthesized barrier by in the the acylation membranes of short-chainof bacteria. 3-hydroxy fatty acids and the addition of ornithine to the free carboxy group of the hydroxy fatty acid. These complex acylated2. FAHFA ornithines Lipid actFamilies as a permeability barrier in the membranes of bacteria. 2. FAHFA2.1. Branched-Chain Lipid Families FAHFA The discovery of branched-chain FAHFAs (Figure 1) was based on their efficacy as endogenous 2.1. Branched-Chain FAHFA anti-diabetic and anti-inflammatory lipids [1]. The anti-diabetic actions appear to involve selective Theagonism discovery at G-protein-coupled of branched-chain receptor40 FAHFAs (GPR40), (Figure thereby1) was basedaugmen onting their glucose e fficacy stimulation as endogenous of anti-diabeticinsulin release and anti-inflammatory [2]. The internal branched lipids [FAHFAs1]. The anti-diabeticare a diverse actionslipid class appear of at least to involve 51 FAHFA selective agonismfamilies at G-protein-coupled and 301 regioisomers receptor40 [3]. Hence, (GPR40), high-resolution thereby augmenting chromatography glucose coupled stimulation with mass of insulin spectrometry is essential to assess the potential roles of different families in physiology [3–5]. release [2]. The internal branched FAHFAs are a diverse lipid class of at least 51 FAHFA families and The most studied FAHFAs are the palmitic-hydroxystearic (PAHSA) and palmitic- 301 regioisomershydroxypalmitic [3]. (PAHPA) Hence, high-resolutionfamilies, which include chromatography the 5-, 7-, 8-, coupled 9-, 10-, 11-, with 12-, mass and spectrometry13-hydroxy is essentialregioiomers to assess [5,6]. the While potential these roles FAHFAs of diff undergoerent families de novo in synthesis physiology in most [3–5 ].organs, they also are Thepresent most in studieddietary plant FAHFAs sources are [7,8]. the palmitic-hydroxystearic Biosynthesis of branched-chain (PAHSA) FAH andFAs palmitic-hydroxypalmitic can be augmented by (PAHPA)oxidative families, stress which[9] and involves include the the transfer 5-, 7-, 8-,of a 9-,fatty 10-, acid 11-, from 12-, a fatty and acid-CoA 13-hydroxy to a regioiomershydroxy fatty [5,6].