Bonds Of Unsaturated Fatty Acids Research Articles

Pulsed Direct Current Arc-Induced Nanoelectrospray Ionization Mass Spectrometry.

This study explores the innovative field of pulsed direct current arc-induced nanoelectrospray ionization mass spectrometry (DCAI-nano-ESI-MS), which utilizes a low-temperature direct current (DC) arc to induce ESI during MS analyses. By employing a 15 kV output voltage, the DCAI-nano-ESI source effectively identifies various biological molecules, including angiotensin II, bradykinin, cytochrome C, and soybean lecithin, showcasing impressive analyte signals and facilitating multicharge MS in positive- and negative-ion modes. Notably, results show that the oxidation of fatty acids using a DC arc produces [M + O - H]- ions, which aid in identifying the location of C═C bonds in unsaturated fatty acids and distinguishing between isomers based on diagnostic ions observed during collision-induced dissociation tandem MS. This study presents an approach for identifying the sn-1 and sn-2 positions in phosphatidylcholine using phosphatidylcholine and nitrate adduct ions, accurately determining phosphatidylcholine molecular configurations via the Paternò-Büchi reaction. With all the advantages above, DCAI-nano-ESI holds significant promise for future analytical and bioanalytical applications.

Fatty acid unsaturation improves germination of upland cotton (Gossypium hirsutum) under cold stress.

The level of fatty acid unsaturation in seeds is one of the major determinants of cold germination ability, particularly in oilseeds. The presence of cis double bonds in unsaturated fatty acids creates bends that lowers their melting temperatures compared to saturated fatty acids. Unsaturated fatty acids with low melting points mobilize faster at low temperatures providing seeds with sufficient energy for germination. To investigate the effects of fatty acid unsaturation on the ability of cotton seeds to germinate under cold conditions, four recombinant inbred lines (RILs) of cotton with unique fatty acid profiles were evaluated using a set of developmental and biochemical assays at 12°C (critically low temperature), 15°C (cardinal minimum temperature) and 30°C (optimum temperature). Furthermore, whole seed lipidome profiling using liquid chromatography with mass spectrometry was done to compare the lipid compositional changes at 12°C and 30°C after imbibing cotton seeds of all the six genotypes for 0 hours, 3 hours and 6 hours. The RILs with higher unsaturation/saturation ratios registered robust germination performance, lower solute leakage, and optimum water uptake rates under cold stress. Imbibition at 30°C for 8 hours before cold exposure significantly improved the germination of cold sensitive genotypes, indicating that the first few hours of water uptake are critical for cold stress. Whole seed lipidome profiling of all the genotypes specifically associated cold germination ability with higher unsaturation levels of phospholipids during early imbibition. The presence of cis double bonds in phospholipids creates kinks that maintain the fluidity of cell membranes under low temperature. Membrane flexibility under cold conditions is essential for facilitating key germination events including membrane organization and respiration. The current results highlight the importance of fatty acid composition in cold germination ability of upland cotton.

Determination of C═C Positions of Unsaturated Fatty Acids in Foods via Ambient Reactive Desorption Ionization with Water Dimer Radical Cations.

The positions of C═C bonds in unsaturated fatty acids (FAs) are one of the main factors determining the quality of food flavor. Herein, we developed an approach for the determination of C═C bonds of FAs by online epoxidation reaction with water dimer radical cations. The limit of detection for octenoic acid isomers was ∼9 μg/L. The positions of C═C bonds in trans-2/3-hexenoic acid, trans-2/3-octenoic acid, oleic acid, linoleic acid, and linolenic acid in black tea or olive oil samples were directly determined by the established method. These results indicate that the established method allows the rapid determination of unsaturated FAs in black tea and olive oil. The advantages of this approach include the analysis speed (∼1 min per sample), simple device, and no need for complex pretreatment. This study not only provides a strategy for the determination of C═C positions but also offers new possibilities for applications in the field of food chemistry.

Detection and analysis of triacylglycerol regioisomers via electron activated dissociation (EAD) tandem mass spectrometry

Triacylglycerols (TGs) are important components of human diet. The positional distribution of fatty acids (FAs) on the glycerol backbone affects the chemistry and physical properties of fats. Especially for infants, the structure of TGs plays an important role in the growth and development. However, limited by detecting technology, accurately identifying regioisomers of ABA/AAB and BAC/ABC/ACB type TGs is a significant challenge for human milk utilization and the development of infant formula. For this, we exploit a novel method for identifying the regioisomers of ABA/AAB and BAC/ABC/ACB type TGs within complex lipid mixtures, via used electron activated dissociation (EAD) tandem mass spectrometry. The distribution information of acyl chains at the sn-2 and sn-1/3 positions of glycerol backbone and double bonds in unsaturated FAs can be easily obtained by fragmenting TG ions with energetic electrons (15 eV). Then, the standard curve was established by correlating the peak area intensity of sn-2 characteristic product ion with the content of TG regioisomers standard. These analytical methods successfully enabled the identification and quantification of TG regioisomers in human milk, cow milk, infant formula, palm oil, and sunflower oil. Additionally, the distribution of the double-bond positions of unsaturated FAs in these samples was also identified. Compared to traditional methods, this approach eliminates the need for complex processing and analysis procedures, enabling rapid structural characterization of ABA/AAB and BAC/ABC/ACB type TGs within 17 min. Hence, we provide a rapid and convenient methodology for detecting and analyzing ABA/AAB and BAC/ABC/ACB type TG regioisomers, thereby offering valuable assistance in the development of specialized formulations and facilitating effective process control for ensuring the quality of edible oils and fats.

Analysis of unsaturated fatty acids by supercritical fluid chromatography tandem mass spectrometry coupled with online Paternò-Büchi reaction

Carbon double bond (CC) positions in unsaturated fatty acids (UFAs) can be identified based on Paternò-Büchi (PB) reaction coupled with mass spectrometry (MS). However, achieving a high conversion rate and confusing mass spectrum are still challenging. In this study, an online PB reaction coupled with supercritical fluid chromatography-mass spectrometry (SFC-MS), using a homemade flow microreactor, was established to analyze CC bonds in UFAs, and benzaldehyde (BA) was proposed as the optimized reagent. Benefiting from this setup, both prototype and product ions of FAs can be rapidly captured at the same retention time, significantly enhancing the accuracy of lipid identification, especially when dealing with complex compounds. Furthermore, a high CC bond conversion rate was achieved, ranging from 53 to 67 %. Additionally, this strategy demonstrated excellent relative quantification abilities, which were based on the ratios of either the sum of diagnostic ions or the individual diagnostic ions. This novel approach was proposed and applied for the specific recognition of free FA in human plasma, which successfully provides quantitative information about UFA isomers. This strategy is believed to hold promise for the analysis of unsaturated lipid isomers in complex biological samples.

Unraveling the Double Bond Position of Fatty Acids by GC-MS Using Electron Capture APCI and In-Source Fragmentation Patterns.

The position of double bonds in unsaturated fatty acids is strongly connected to their biological effects, but their analytical characterization is still challenging. However, the ionization of unsaturated fatty acids by a GC-APCI leads to regiospecific in-source fragment ions, which can be used to identify the double bond position. The fragment ions are oxidized species that occur mostly at the double bond closest to the carboxylic acid group. This effect can be further promoted by using benzaldehyde as a gas-phase reactant. This allows the identification of the Δ-notation of the fatty acid, and based on additional information such as m/z and retention time, it is possible to annotate the corresponding fatty acid. The developed method also enables the quantification of fatty acids in one step with high selectivity and sensitivity. Moreover, rare fatty acids can be identified in suspected target approaches that are often not available as standards. This was demonstrated by analyzing fish oil samples that provide a complex mixture of highly unsaturated fatty acids and by identifying rare fatty acids such as hexadecatetraenoic acid (FA 16:4 Δ6).

Simultaneous multiplexed quantification and C=C localization of fatty acids with LC-MS/MS using isobaric multiplex reagents for carbonyl-containing compound (SUGAR) tags and C=C epoxidation

Fatty acids (FAs) possess highly diverse structures and can be divided into saturated and unsaturated classes. For unsaturated FAs, both the numbers and positions of carbon-carbon double bond (C=C) determine their biological functions. Abnormal levels of FA isomers have been reported to be involved in various disease development, such as cancer. Despite numerous advances in lipidomics, simultaneous quantifying and pinpointing C=C bond positions in a high-throughput manner remains a challenge. Here we conducted epoxidation of C=C bonds of unsaturated FAs followed by the conjugation with isobaric SUGAR tags. With the assistance of LC-MS, FA isomers with the same masses were separated on the C18 column and individually subjected to MS/MS fragmentation. Upon higher-energy collisional dissociation, not only reporter ions for multiplexed quantification but also diagnostic ions for C=C localization were generated at the same time, allowing quantitative analyses of different unsaturated FA isomers in samples. The performance of this approach including epoxidation, labeling efficiencies, quantitation accuracy, and capability to pinpoint C=C bond position were evaluated. To evaluate our method, free FA extracts from healthy human serum were used to demonstrate the feasibility of this method for complex sample analysis. Finally, this method was also applied to investigate the changes of unsaturated FA isomers between heathy human and Alzheimer's disease (AD) patient serum.

Reactive Thread Spray Mass Spectrometry for Localization of C═C Bonds in Free Fatty Acids: Applications for Obesity Diagnosis.

Cellulose thread substrates offer a platform for microsampling and reactive ionization of free fatty acid (FFA) isomers for direct differentiation by mass spectrometry. Ambient corona discharge forms when direct current high voltage is applied to the tiny subfibers on the thread substrate in the presence of a polar spray solvent (MeOH/H2O, 2:1, v/v), facilitating chemical reactions across a C═C bond of unsaturated fatty acids. The process was applied for diagnosis of obesity, which we observed to show better discriminatory power when compared to determinations based on body mass index. Overall, the integrated reactive thread-based platform is capable of (i) microsampling and dry-state, room-temperature storage (>30 days) of the biofluids, (ii) in-capillary liquid/liquid extraction, and (iii) in situ epoxidation reactions to locate the C═C bond position in unsaturated fatty acids via reactions with reactive oxygen species present in ambient corona discharge. The study showcased the ability to correctly characterize FFAs, including degree of unsaturation, and the determination of their relative concentrations in clinical biofluid samples.

Palm oil and cinnamon (anti-microbial agent) on the physicochemical, mechanical, and biodegradation properties of micrometer-sized cornstarch-based bioplastic

The purpose of this study was to determine the effect of the addition of palm oil and cinnamon on the physicochemical, mechanical, and biodegradation properties of cornstarch-based bioplastic. Experiments were done by adding cornstarch into water, glycerol, acetic acid, and cinnamon. Then, the mixed suspension was heated at 100⁰C, added with palm oil (0.19; 0.30; 0.60; 0.90; 1.30; 1.90 %(v/v)), molded, and dried at room temperature without exposure to sunlight. To support the analysis, several characterizations (i.e. FTIR, digital microscope, puncture test, compressive test, and biodegradability test) were conducted. Based on puncture and compressive test, the bioplastic with an addition of 1.90 %(v/v) palm oil showed good mechanical properties. In general, the addition of palm oil increased the biodegradation rate. The palm oil contains double bonds in unsaturated fatty acids that can be distributed to the bioplastic matrix. Higher lipid content caused less moisture in the bioplastic so that the bioplastic was more brittle, inelastic, and degradable. However, the antimicrobial amount in the cinnamon reduced the biodegradability of bioplastic. This research provided new information on bioplastics as environmentally friendly plastic and alternative to conventional plastic.

Non-covalent double bond sensors for gas-phase infrared spectroscopy of unsaturated fatty acids

The position and configuration of carbon-carbon double bonds in unsaturated fatty acids is crucial for their biological functions and influences health and disease. However, double bond isomers are not routinely distinguished by classical mass spectrometry workflows. Instead, they require sophisticated analytical approaches usually based on chemical derivatization and/or instrument modification. In this work, a novel strategy to investigate fatty acid double bond isomers (18:1) without prior chemical treatment or modification of the ion source was implemented by non-covalent adduct formation in the gas phase. Fatty acid adducts with sodium, pyridinium, trimethylammonium, dimethylammonium, and ammonium cations were characterized by a combination of cryogenic gas-phase infrared spectroscopy, ion mobility-mass spectrometry, and computational modeling. The results reveal subtle differences between double bond isomers and confirm three-dimensional geometries constrained by non-covalent ion-molecule interactions. Overall, this study on fatty acid adducts in the gas phase explores new avenues for the distinction of lipid double bond isomers and paves the way for further investigations of coordinating cations to increase resolution.Graphical abstract

Location of carbon-carbon double bonds in unsaturated lipids using microdroplet mass spectrometry.

An aqueous solution containing unsaturated fatty acids (100 μM) or lipids (50 μg mL-1) and chloroauric acid (HAuCl4, 10 μM) is electrosprayed (-4.5 kV for unsaturated fatty acids and +4.0 kV for lipids) from a 50 μm diameter capillary with N2 nebulizing gas (60 psi), and the resulting microdroplets enter a mass spectrometer with a flight distance of 10 mm for chemical analysis. The HAuCl4 oxidizes the C[double bond, length as m-dash]C double bond to cause the formation of an aldehyde group or a hydroxyl group on one side and a carboxyl group on the other (i.e., CHO-R-COOH or HO-R-COOH), allowing the location of the double bond to be identified. This approach was successfully applied to four unsaturated fatty acids [linoleic acid (LA), ricinoleic acid (RA), isooleic acid (IA), and nervonic acid (NA)] and two phospholipids [1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) and L-α-lysophosphatidylcholine (lysoPC)]. A mechanism for this transformation is proposed, which involves epoxidation of the double bond, followed by the formation of the final products. This method has the advantages of being simple and rapid, and requiring a small amount of analyte.

Positional determination of the carbon\u2013carbon double bonds in unsaturated fatty acids mediated by solvent plasmatization using LC\u2013MS

Fatty acids (FAs) are the central components of life: they constitute biological membranes in the form of lipid, act as signaling molecules, and are used as energy sources. FAs are classified according to their chain lengths and the number and position of carbon–carbon double bond, and their physiological character is largely defined by these structural properties. Determination of the precise structural properties is crucial for characterizing FAs, but pinpointing the exact position of carbon–carbon double bond in FA molecules is challenging. Herein, a new analytical method is reported for determining the double bond position of mono- and poly-unsaturated FAs using liquid chromatography-mass spectrometry (LC–MS) coupled with solvent plasmatization. With the aid of plasma on ESI capillary, epoxidation or peroxidation of carbon–carbon double bond in FAs is facilitated. Subsequently, molecular fragmentation occurs at or beside the epoxidized or peroxidized double bond via collision-induced dissociation (CID), and the position of the double bond is elucidated. In this method, FAs are separated by LC, modified by plasma, fragmented via CID, and detected using a time-of-flight mass spectrometer in a seamless manner such that the FA composition in a mixture can be determined. Our method enables thorough characterization of FA species by distinguishing multiple isomers, and therefore can uncover the true diversity of FAs for their application in food, health, and medical sciences.

Derivatisation of 2,4 (dinitrophenyl hydrazine) DNPH in Canola oil oxidation

Lipid oxidation during heating started at the allylic position of the double bond of unsaturated fatty acids. Primary lipid oxidation which was produced during heating are generally unstable and directly decompose into secondary oxidation products. The oxidative and thermal stability of Rapeseed oil was evaluated using Rancimat 679. Oil samples were heated at 120oC for 10 hours which are taken started from 5, 6, 6.5, 7, 7.5, 8, 9 and 10 hours. Derivatization with DNPH eluted with a mixture of methanol (60%) and water (40%). The aldehydes are analysed by High Performance Liquid Chromatography- Diode Array Detection (HPLC-DAD) after derivatisation with DNPH and identified by high-resolution MS using an orbitrap-based mass spectrometer. HPLC is the suitable method to investigate the non-volatile aldehydes formed by decomposition of hydroperoxides. The result of this oxidation process were described that the maximum point of the oil oxidation was at 9 hours which peak area 378. For oxidation process at the 10 hours the area peak is decreased which point 344.

Determination and quantification of fatty acid C=C isomers by epoxidation reaction and liquid chromatography-mass spectrometry

The location of double bond in unsaturated fatty acids (FAs) plays a critical role in their physiological properties. However, structural identification and quantification of unsaturated FAs by mass spectrometry are still challenging. In this work, we reported the coupling of epoxidation reaction of the C=C in unsaturated FAs and liquid chromatography-mass spectrometry (LC-MS) with multiple reaction monitoring (MRM) mode for accurate identification and quantification of C=C isomers of FAs. Epoxidation of the C=C in unsaturated FAs was induced by a dioxide of ketone, tetrahydrothiopyran-4-one 1,1-dioxide, as a catalyst and Oxone as an oxidant in less than 5 min with nearly 100% yield. All the C=C bonds were epoxidized to obtain a single product, simplifying the chromatographic separation of epoxidation products to enable more accurate quantification analysis. The epoxidation products were stable at room temperature and can produce highly abundant diagnostic ions indicative of C=C locations by tandem mass spectrometry using collision-induced association (CID). The application of this approach for the analysis of FAs isomers in human plasma demonstrated the potential of our method for the qualitative and quantitative analysis of unsaturated FAs in complex biological samples, which is valuable in biological and medical analysis.

Characterization of different ozonized sunflower oils II. Triacylglycerol condensation and physical properties

Within the chemical transformations that vegetable oils undergo, oxidative reactions are one of the most widely studied. During the process of oxidation oils react with oxygen in their different forms to produce a large variety of chemical species that alter their chemical and physical properties. This work is the second of a series of two in which the action of ozone on different sunflower oils was investigated. Ozone reacts with double bonds of unsaturated fatty acids which produce different peroxidated species. The presence of altered triacylglycerol (TAG) increases the polarity of the vegetable oils and induces changes in their physical properties. Moreover, peroxygenation alters the aggregation of TAGs, favoring the formation of dimers or oligomers that can substantially change the melting and crystallization profiles of these oils. In the present work the effect of ozonation on four different sunflower oils was studied: common sunflower, high oleic sunflower and two highly saturated sunflower oils, high palmitic and stearic in a high oleic background. Furthermore, the species of TAGs from the different oils that were affected by the ozone oxidative attack were studied. The formation of polar compounds and TAG aggregates were investigated as well as the effect caused by them on the physical properties of the oils as studied through differential scanning calorimetry.

Preparation and Characterization of PEGylated C18 Fatty Acids/Anti-SNAP25 Antibody-Targeted Liposomes

Background: Unsaturated C18 fatty acids, such as oleic acid (L1), linoleic acid (L2), and linolenic acid (L3), are a good choice of lipids to prepare liposomes. They are inexpensive, biocompatible, nontoxic, and readily available compared with phospholipids. Moreover, cis-double bonds of unsaturated fatty acids prevent the packing of molecules which increases membrane fluidity in liposomes making them a good choice of starting materials to prepare liposomes. Objective: Unsaturated C18 fatty acid liposomes, as well as their PEGylated and non- PEGylated antibody-targeted liposomes, were prepared and characterized. Methods: The particle size and zeta potential of the prepared liposomes (1 mM, pH = 7.4) for 28 and 14 days, respectively, were monitored and characterized. Membrane-bound antibodies Anti-SNAP25 (AS25) and DOPE PEG2000 (DP) were conjugated to pure C18 fatty acid liposomes to achieve stable fatty acid formulations. Results: The mean particle sizes of pure L1, L2, and L3 liposome solutions were 125, 129, and 122 nm respectively, while their polydispersity index values were 0.28, 0.21, and 0.40 respectively. A large negative zeta potential value of 45 mV was observed due to anionic carboxylate head-group of pure liposomes. The incorporation of AS25 into L1/DP, L2/DP, and L3/DP liposome solutions stabilized their mean particle size and zeta potential measurements over 28 and 14 days, respectively. Conclusion: L1/DP/AS25 was found to be the most stable PEGylated antibody-targeted liposome system because its particle size remained between 90 and 125 nm in 28 days. Transmission electron microscopy observations also supported the incorporation of AS25 and DP on the membrane surface as predicted.

Oleate hydratase from Staphylococcus aureus protects against palmitoleic acid, the major antimicrobial fatty acid produced by mammalian skin

Oleate hydratases (OhyAs) belong to a large family of bacterial proteins catalyzing the hydration or isomerization of double bonds in unsaturated fatty acids. A Staphylococcus aureus gene (Sa0102) is predicted to encode an OhyA. Here, we recombinantly expressed and purified SaOhyA and found that it forms a homodimer that requires FAD for activity. SaOhyA hydrates only unsaturated fatty acids containing cis-9 double bonds, but not fatty acids with trans-9 double bonds or cis double bonds at other positions. SaOhyA products were not detected in S. aureus phospholipids and were released into the growth medium. S. aureus does not synthesize unsaturated fatty acids, and the SaOhyA substrates are derived from infection sites. Palmitoleate (16:1(9Z)) is a major mammalian skin-produced antimicrobial fatty acid that protects against S. aureus infection, and we observed that it is an SaOhyA substrate and that its hydroxylated derivative is not antimicrobial. Treatment of S. aureus with 24 μm 16:1(9Z) immediately arrested growth, followed by growth resumption after a lag period of 2 h. The ΔohyA mutant strain did not recover from the 16:1(9Z) challenge, and increasing SaOhyA expression using a plasmid system prevented the initial growth arrest. Challenging S. aureus with sapienic acid (16:1(6Z)), an antimicrobial fatty acid produced only by human skin, arrested growth without recovery in WT, ΔohyA, and SaOhyA-overexpressing strains. We conclude that SaOhyA protects S. aureus from palmitoleic acid, the antimicrobial unsaturated fatty acid produced by most mammals, and that sapienic acid, uniquely produced by humans, counters the OhyA-dependent bacterial defense mechanism.

Effect of dietary olive oil and palm oil on rumen bacterial composition in dairy cows

The rumen bacteria play a major role in lipid metabolism. Bacteria remove the double bonds of unsaturated fatty acids resulting in the production of saturated fatty acids, which are incorporated in milk. Crude olive oil (rich in unsaturated fatty acids) represents a potentially valuable feed source for dairy cows that might enhance the human-health beneficial composition of milk and dairy products. This project studied the effect of supplementing dairy cow diets with olive oil (OO) and palm oil (HVO) on rumen microbiota. For 63 days the animals were fed a control diet (basal diet) with no added lipid and two fat-supplemented diets (30 g kg−1 DM). Rumen sampling were performed at the onset of the experiment and every 21 days for 63 days using a rumen scoop. Total microbial DNA was extracted from ruminal samples for high-throughput sequencing of the 16S rRNA gene through Illumina MiSeq platform. Results revealed the dominance of phyla Firmicutes and Bacteroidetes. Firmicutes was the most prevalent phyla in diet control (75.2 %), OO (71.1 %) and HVO (75.2 %). At genus level Succiniclasticum and Prevotella were the dominant genera, belonging to Firmicutes and Bacteroidetes respectively. Succiniclasticum decrease significantly their relative abundance during OO supplementation (p)

Double bonds of unsaturated fatty acids differentially regulate mitochondrial cardiolipin remodeling

BackgroundSupplemented fatty acids can incorporate into cardiolipin (CL) and affect its remodeling. The change in CL species may alter the mitochondrial membrane composition, potentially disturbing the mitochondrial structure and function during inflammation.MethodTo investigate the effect of the unsaturation of fatty acids on CL, we supplemented macrophage-like RAW264.7 cells with 18-carbon unsaturated fatty acids including oleic acid (OA, 18:1), linoleic acid (LA, 18:2), α-linolenic acid (ALA, 18:3), γ-linolenic acid (GLA, 18:3), and stearidonic acid (SDA, 18:4). Mitochondrial changes in CL were measured through mass spectrometry.ResultOur data indicated that OA(18:1) was the most efficient fatty acid that incorporated into CL, forming symmetrical CL without fatty acid elongation and desaturation. In addition, LA(18:2) and ALA(18:3) were further elongated before incorporation, significantly increasing the number of double bonds and the chain length of CL. GLA and SDA were not optimal substrates for remodeling enzymes. The findings of RT-qPCR experiments revealed that none of these changes in CL occurred through the regulation of CL remodeling- or synthesis-related genes. The fatty acid desaturase and transportation genes—Fads2 and Cpt1a, respectively—were differentially regulated by the supplementation of five unsaturated 18-carbon fatty acids.ConclusionsThe process of fatty acid incorporation to CL was regulated by the fatty acid desaturation and transportation into mitochondria in macrophage. The double bonds of fatty acids significantly affect the incorporation process and preference. Intact OA(18:1) was incorporated to CL; LA(18:2) and ALA(18:3) were desaturated and elongated to long chain fatty acid before the incorporation; GLA(18:3) and SDA(18:4) were unfavorable for the CL incorporation.

Sequence-Defined Cationic Lipo-Oligomers Containing Unsaturated Fatty Acids for Transfection.

Sequence-defined cationic lipo-oligomers containing unsaturated fatty acids are potent nucleic acid carriers that are produced by solid-phase supported synthesis. However, the trifluoroacetic acid (TFA)-mediated removal of acid-labile protecting groups and cleavage from the resin can be accompanied by side products caused by an addition of TFA to the double bonds of unsaturated fatty acids. These TFA adducts are converted into hydroxylated derivatives under aqueous conditions. Here we describe an optimized cleavage protocol (precooling cleavage solution to 4°C, 20min cleavage at 22°C), which minimizes TFA adduct formation, retains the unsaturated hydrocarbon chain character, and ensures high yields of the synthesis.