Fluorescence with fatty acids

Used for the precolumn preparation of fluorescent derivatives of carboxylic acids reagent reacts well with fatty acids at room temperature to give intensely fluorescent esters Reference 2... 

Richieri,G.V.,Ogata,R.T.,andKleinfeld,A.M., 1994,Equilibrium constants for the binding of fatty acids with fatty acid-binding proteins from adipocyte, intestine, heart and liver measured with the fluorescent probe ADIFAB, / Biol. Chem. 269 23918-23930. 

Nimura and Kinoshita [96] developed a derivatization technique for the analysis of fatty acids that is based on 9-anthryldiazomethane (ADAM). The latter is very stable in solution and reacts with fatty acids at room temperature without a catalyst to yield strongly fluorescent esters, which can be chromatographed on an ODS phase with acetonitrile/water mixtures. The derivatization of fatty acid is carried out with a 0.1% methanohc solution of 9-anthryldiazomethane, which is prepared via oxidation of 9-anthraldehyde-hydrazone following a procedure by Nakaya et al. [97]. The fluorescence and excitation spectra of fatty acid-methyl-anthracene esters exhibit maxima at 412 and 365 nm, respectively. As an example, Figure 8.62 shows the separation of ADAM derivatives, a mixture of saturated and unsaturated long-chain fatty acids, which can be detected down to the lowest picomole range. 

Barbana C, Perez MD, Sanchez L, ei al. 2006. Interaction of bovine a-lactalbumin with fatty acids as determined by partition equilibrium and fluorescence spectroscopy. Int Dairy J 16 18-25. 

Acids can also be converted to fluorescent dansyl derivatives The reaction of Cg to C24 fatty acids with dansyl semipiperazide or semicadavende provides an excellent example (Fig 34) [87] Odd-numbered and unsaturated fatty acids [88] and propionic, sorbic and benzoic acid [89] can be detected in the same manner... 

Fig. 1 Fluorescence scan of a fatty acid mixture with 500 ng substance per chromatogram zone. Arachidic acid (I), stearic acid (2), palmitic acid (3), myristic acid (4), lauric acid (5).

Fatty Acid Transporters. Figure 2 Quencher-based real-time fatty acid uptake assay with a fluorescently labeled FFA analogue (C1-Bodipy-C12). Predominantly protein-mediated fatty acid uptake by 3T3-L1 adipocytes (diamonds) was compared with diffusion-driven uptake by fibroblasts (squares) using the QBT Fatty Acid Uptake reagent (Molecular Devices Corp., CA, USA), which contains C1-Bodipy-C12 as substrate in conjunction with a cell impermeable quencher. Uptake kinetics was recorded using a Gemini fluorescence plate reader. Error bars indicate the standard deviations from 12 independent wells. RFU relative fluorescence units. 

The photobleaching of P-carotene by fluorescent light in fatty acid ester solutions showed an autoxidation kinetic profile with the rate of degradation of P-carotene in the order laurate > oleate > linoleate (Carnevale et al. 1979). The presence of a radical scavenger retarded the autoxidation, thus leading to the view that protection against autoxidation is built into the system by the unsaturation in the fatty acid. 

M. Vincent, B. de Foresta, J. Gallay, and A. Alfsen, Nanosecond fluorescence anisotropy decays of n-(9-anthroyloxy) fatty acids in dipalmitoylphosphatidylcholine vesicles with regard to isotropic solvents, Biochemistry 21, 708-716 (1982). 

D. B. Chalpin and A. M. Kleinfeld, Interaction of fluorescence quenchers with the -(9-anthroyloxy) fatty acid membrane probes, Biochim. Biophys. Acta 731, 465 174 (1983). 

Dillard, C. J., and A. L. Tappel. Fluorescent products from reaction of peroxidiz-ing polyunsaturated fatty acids with phosphatidyl ethanolamine and phenylalanine. Upids 8 183-189, 1973. 

Glutathione S transferases bind bile acids in vitro but doubt has been cast over whether this happens in vivo as these enzymes were not labelled by fluorescently labelled bile acids in experiments to identify the carrier proteins but may play a role with the raised levels in cholestasis. Liver fatty-acid-binding protein has been shown to bind bile acids by using a displacement assay with fluorescent fatty-acid ligand. This work clearly showed displacement to be directly related to hydrophobicity, such that lithocholate conjugates had the greatest effect. This may indicate a mechanism to minimise toxicity within the hepatocyte. 

Let us recall the micellar aqueous system, as this procedure is actually the basic one. The chemistry is based on fatty acids, that build micelles in higher pH ranges and vesicles at pH c. 8.0-8.5 (Hargreaves and Deamer, 1978a). The interest in fatty acids lies also in the fact that they are considered possible candidates for the first prebiotic membranes, as will be seen later on. The experimental apparatus is particularly simple, also a reminder of a possible prebiotic situation the water-insoluble ethyl caprylate is overlaid on an aqueous alkaline solution, so that at the macroscopic interphase there is an hydrolysis reaction that produces caprylate ions. The reaction is very slow, as shown in Figure 7.15, but eventually the critical micelle concentration (cmc) is reached in solution, and thus the first caprylate micelles are formed. Aqueous micelles can actually be seen as lipophylic spherical surfaces, to which the lipophylic ethyl caprylate (EC) avidly binds. The efficient molecular dispersion of EC on the micellar surface speeds up its hydrolysis, (a kind of physical micellar catalysis) and caprylate ions are rapidly formed. This results in the formation of more micelles. However, more micelles determine more binding of the water-insoluble EC, with the formation of more and more micelles a typical autocatalytic behavior. The increase in micelle population was directly monitored by fluorescence quenching techniques, as already used in the case of the... 

Similar dicholoro precursor 205 was utilized by Katoh and co-workers in the discovery of a new fluorescence derivatizing agent for fatty acids <99H299>. The conversion of 205 to fatty acid-coupled 206 proceeded through the displacement of both chloro groups with morpholine in several high-yielding steps. 

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