Acylcarnitines

FIGURE 24.9 The formation of acylcar-nitines and their transport across the inner mitochondrial membrane. The process involves the coordinated actions of carnitine acyltrans-ferases on both sides of the membrane and of a translocase that shuttles O-acylcarnitines across the membrane. 

Several additional points should be made. First, although oxygen esters usually have lower group-transfer potentials than thiol esters, the O—acyl bonds in acylcarnitines have high group-transfer potentials, and the transesterification reactions mediated by the acyl transferases have equilibrium constants close to 1. Second, note that eukaryotic cells maintain separate pools of CoA in the mitochondria and in the cytosol. The cytosolic pool is utilized principally in fatty acid biosynthesis (Chapter 25), and the mitochondrial pool is important in the oxidation of fatty acids and pyruvate, as well as some amino acids. 

FIGURE 25.16 Regulation of fatty acid synthesis and fatty acid oxidation are conpled as shown. Malonyl-CoA, produced during fatty acid synthesis, inhibits the uptake of fatty acylcarnitine (and thus fatty acid oxidation) by mitochondria. When fatty acyl CoA levels rise, fatty acid synthesis is inhibited and fatty acid oxidation activity increases. Rising citrate levels (which reflect an abundance of acetyl-CoA) similarly signal the initiation of fatty acid synthesis. 

Figure 3. Mitochondrial fatty acid oxidation. Long-chain fatty acids are converted to their CoA-esters as described in the text, and their fatty-acyl-groups transferred to CoA in the matrix by the concerted action of CPT 1, the acylcarnitine/carnitine exchange carrier and CPT (A) as described in the text. Medium-chain and short-chain fatty acids (Cg or less) diffuse directly into the matrix where they are converted to their acyl-CoA esters by a acyl-CoA synthase. The mechanism of p-oxidation is shown below (B). Each cycle of P-oxidation removes -CH2-CH2- as an acetyl unit until the fatty acids are completely converted to acetyl-CoA. The enzymes catalyzing each stage of P-oxidation have different but overlapping specificities. In muscle mitochondria, most acetyl-CoA is oxidized to CO2 and H2O by the citrate cycle (Figure 4) some is converted to acylcamitine by carnitine acetyltransferase (associated with the inner face of the inner membrane) and exported from the matrix. Some acetyl-CoA (if in excess) is hydrolyzed to acetate and CoASH by acetyl-CoA hydrolase in the matrix. Enzymes ...

Carnitine (p-hydroxy-y-trimethylammonium butyrate), (CHjljN"—CH2—CH(OH)—CH2—COO , is widely distributed and is particularly abundant in muscle. Long-chain acyl-CoA (or FFA) will not penetrate the inner membrane of mitochondria. However, carnitine palmitoyltransferase-I, present in the outer mitochondrial membrane, converts long-chain acyl-CoA to acylcarnitine, which is able to penetrate the inner membrane and gain access to the P-oxidation system of enzymes (Figure 22-1). Carnitine-acylcar-nitine translocase acts as an inner membrane exchange transporter. Acylcarnitine is transported in, coupled with the transport out of one molecule of carnitine. The acylcarnitine then reacts with CoA, cat-... 

Figure 22-1. Role of carnitine in the transport of long-chain fatty acids through the inner mitochondrial membrane. Long-chain acyl-CoA cannot pass through the inner mitochondrial membrane, but its metabolic product, acylcarnitine, can. 

A few patients have been described with a defect involving the carnitine-acylcarnitine translocase system, which facilitates the movement of long-chain acylcarnitine esters across the inner membrane of the mitochondrion (Fig. 42-2). These patients have extremely low carnitine concentrations and minimal dicarboxylic aciduria [4]. 

This in fact is what happened. Tandem MS has clearly been shown to be the only technology to screen for disorders of fatty acid oxidation and could also detect many disorders of organic acid metabolism. Tandem MS has the ability to detect both compound classes (amino acids and acylcarnitines) and after demonstrating that both classes could be prepared in the same manner, the MS/MS analysis of blood spots for newborn screening applications was born. 

Figure 14.2. Acylcarnitine profile obtained using a precursor ion scan of 85 Da. The profile is from the blood spot of a normal patient.

Esterase ESI5 Acylcarnitine hydrolase Carboxylesterase RL2 Esterase pi 5.5 Hydrolase pi 5.6 Acylcarnitine hydrolase Esterase Esterase I Esterase ES10 Carboxylesterase RH1 Octanoylglycerol lipase Esterase pi 6.1 Hydrolase pi 6.2... 

R. Mentlein, H. Rix-Matzen, E. Heymann, Subcellular Localization of Nonspecific Carboxylesterases, Acylcarnitine Hydrolase, Monoacylglycerol Lipase and Palmitoyl-CoA Hydrolase in Rat Liver , Biochim. Biophys. Acta 1988, 964, 319-328. 

A simple and efficient alternative to the traditional UV detection of amino acids and related compounds is nowadays represented by the evaporative light scattering (ELS) detector, which allows the direct chromatographic separation, with no need for preliminary derivatization. In the field of glycopeptides-based CSPs, it was applied for the first time in the chromatographic resolution of carnitine and 0-acylcarnitine enantiomers on a TE CSP [61]. The considered compounds are nonvolatile solids and gave optimal ELS response under a variety of experimental conditions (buffered and unbuffered mobile phases, flow-rates from 0.5 to 1.5 mL/min, different kind and... 

Carnitine is a vitamin-like quaternary ammonium salt, playing an important role in the human energy metabolism by facilitating the transport of long-chained fatty acids across the mitochondrial membranes. An easy, fast, and convenient procedure for the separation of the enantiomers of carnitine and 0-acylcarnitines has been reported on a lab-made teicoplanin-containing CSP [61]. The enantioresolution of carnitine and acetyl carnitine was enhanced when tested on a TAG CSP, prepared in an identical way [45]. Higher a values were reached also in the case of A-40,926 CSP [41]. 

D Acquarica, I. et al., Direct chromatographic resolution of carnitine and O-acylcarnitine enantiomers on a teicoplanin-bonded chiral stationary phase,... 

Heinig, K., and Henion, J. (1999). Determination of carnitine and acylcarnitines in biological samples by capillary electrophoresis-mass spectrometry.. Chromatogr. B 735, 171 — 188. 

Fatty acyl carnitine is transported via a translocase that transports acylcarnitine into and carnitine out of the mitochondrion (Chapter 7). 

CH3)3N+-CH2-CH(0H)-CH2-C0Q-, an acyl-transfer metabolite, found in high abundance in skeletal muscle, liver, and yeast. O-Acylcarnitine, is a high-energy ester like its structurally related O-acetylcholine, (CH3)3N+ - CH2 - CH2 - O - C(=O) - CH3. 

Selected entries from Methods in Enzymology [vol, page(s)] Radioisotopic assay of acetylcarnitine and acetyl-CoA, 123, 259 short-chain acylcarnitines identification and quantitation, 123,... 

Tonazzi, A., Giangregorio, N., Indiveri, C. and Palmieri, F. 2005. Identification by site-directed mutagenesis and chemical modification of three vicinal cysteine residues in rat mitochondrial camitine/acylcarnitine transporter. Journal of Biological Chemistry, 280 19607-19612. 

Pharmacological approaches include the inhibition of release of arachidonic acid by inhibition of phospholipase A2 and the inhibition of acylcarnitine transferase I by and oxfenicine, the latter of which has been shown to prevent or at least delay ischemia-induced uncoupling. There are at present no data available on the possible effects of inhibitors of arachidonic acid release on ischemia-induced uncoupling. 

Fischbach PS, Corr PB, Yamada KA Long-chain acylcarnitine increases intracellular calcium and induces afterdepolarizations in adult ventricular myocytes (abstracts). Circulation 1992 86(suppl I) 748. 

Wu J, Corr PB Influence of long chain acylcarnitines on the voltage-dependent calcium current in adult ventricular myocytes. Am J Phyisol 1992 263 H410-H417. 

Wu J, McHowat J, Saffitz JE, Yamada KA, Corr PB Inhibition of gap junctional conductance by long chain acylcarnitines and their preferential accumulation in junctional sarcolemma during hypoxia. Circ Res 1993 72 879-889. 

Acylcarnitine analysis for the diagnosis of organic acidemias and particularly of FAO disorders plays an increasingly prominent role in all venues of clinical biochemical genetics prenatal diagnosis, newborn screening, evaluation of symptomatic patients, and postmortem screening. Almost exclusively performed by tandem... 

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