Aspects of carnitine ester metabolism in sheep liver

1. Carnitine acetyltransferase (EC 2.3.1.7) activity in sheep liver mitochondria was 76nmol/min per mg of protein, in contrast with 1.7 for rat liver mitochondria. The activity in bovine liver mitochondria was comparable with that of sheep liver mitochondria. Carnitine palmitoyltransferase activity was the same in both sheep and rat liver mitochondria. 2. The [free carnitine]/[acetylcarnitine] ratio in sheep liver ranged from 6:1 for animals fed ad libitum on lucerne to approx. 1:1 for animals grazed on open pastures. This change in ratio appeared to reflect the ratio of propionic acid to acetic acid produced in the rumen of the sheep under the two dietary conditions. 3. In sheep starved for 7 days the [free carnitine]/[acetylcarnitine] ratio in the liver was 0.46:1. The increase in acetylcarnitine on starvation was not at the expense of free carnitine, as the amounts of free carnitine and total acid-soluble carnitine rose approximately fivefold on starvation. An even more dramatic increase in total acid-soluble carnitine of the liver was seen in an alloxan-diabetic sheep. 4. The [free CoA]/[acetyl-CoA] ratio in the liver ranged from 1:1 in the sheep fed on lucerne to 0.34:1 for animals starved for 7 days. 5. The importance of carnitine acetyltransferase in sheep liver and its role in relieving ;acetyl pressure' on the CoA system is discussed.     

Effects of ciprofibrate and 2-[5-(4-chlorophenyl)pentyl]oxirane-2-carboxylate (POCA) on the distribution of carnitine and CoA and their acyl-esters and on enzyme activities in rats. Relation between hepatic carnitine concentration and carnitine acetyltransferase activity.

The effects of feeding the peroxisome proliferators ciprofibrate (a hypolipidaemic analogue of clofibrate) or POCA (2-[5-(4-chlorophenyl)pentyl]oxirane-2-carboxylate) (an inhibitor of CPT I) to rats for 5 days on the distribution of carnitine and acylcarnitine esters between liver, plasma and muscle and on hepatic CoA concentrations (free and acylated) and activities of carnitine acetyltransferase and acyl-CoA hydrolases were determined. Ciprofibrate and POCA increased hepatic [total CoA] by 2 and 2.5 times respectively, and [total carnitine] by 4.4 and 1.9 times respectively, but decreased plasma [carnitine] by 36-46%. POCA had no effect on either urinary excretion of acylcarnitine esters or [acylcarnitine] in skeletal muscle. By contrast, ciprofibrate decreased [acylcarnitine] and [total carnitine] in muscle. In liver, ciprofibrate increased the [carnitine]/[CoA] ratio and caused a larger increase in [acylcarnitine] (7-fold) than in [carnitine] (4-fold), thereby increasing the [short-chain acylcarnitine]/[carnitine] ratio. POCA did not affect the [carnitine]/[CoA] and the [short-chain acylcarnitine]/[carnitine] ratios, but it decreased the [long-chain acylcarnitine]/[carnitine] ratio. Ciprofibrate and POCA increased the activities of acyl-CoA hydrolases, and carnitine acetyltransferase activity was increased 28-fold and 6-fold by ciprofibrate and POCA respectively. In cultures of hepatocytes, ciprofibrate caused similar changes in enzyme activity to those observed in vivo, although [carnitine] decreased with time. The results suggest that: (1) the reactions catalysed by the short-chain carnitine acyltransferases, but not by the carnitine palmitoyltransferases, are near equilibrium in liver both before and after modification of metabolism by administration of ciprofibrate or POCA; (2) the increase in hepatic [carnitine] after ciprofibrate or POCA feeding can be explained by redistribution of carnitine between tissues; (3) the activity of carnitine acetyltransferase and [total carnitine] in liver are closely related.     

Release of carnitine from the perfused rat liver

Perfused rat liver was shown to be the proper model for studies on hepatic cellular transport of carnitine. During recirculating perfusion the livers kept equilibrium with 45 nmol/ml total carnitine in perfusate, exhibited concentrative uptake and there was no sign of artificial leakage. The release side of the carnitine transport was characterized by utilizing outflow perfusions. The livers from fed rats exported daily 9.93 mumol per 100 g body weight total carnitine. This release rate is 4- or 10-fold higher than the estimated daily turnover in vivo or the measured urinary excretion. Therefore, the major part of the released carnitine has to re-enter the liver. The outward carnitine transport does not depend on energy or the Na+-K+ pump, since it did not respond to metabolic poisons and ouabain. However, the release rate was strongly inhibited by mersalyl and showed saturability in function of tissue carnitine levels. The Vmax of the saturable outward transport system was 2.47 nmol . min-1 . g-1 liver, the apparent Km was 0.27 mM tissue level (both as compared to total carnitine). These data showed the outward transport of carnitine from the liver to be protein mediated. The contribution of a diffusion (nonsaturable) component was estimated to be 20-25% in the range of tissue levels occurring in vivo. The rate of carnitine release from the liver decreased as an effect of 24 h starvation from the daily 9.92 mumol release to 6.55 mumol on 100 g body weight basis. This decrease is more pronounced when the release rates are expressed on the basis of tissue carnitine levels. The resulting value can be called rate constant (at the linear part of the saturation curve, Fig. 5) and it decreased to 5.00 min-1 from 8.41 min-1 as an effect of starvation. We have concluded that the altered parameters of carnitine transport across the liver cell is decisive in developing the higher hepatic carnitine concentration in the fasted state.     

Metabolism of carnitine in phenylacetic acid-treated rats and in patients with phenylketonuria

The effect of metabolites accumulating in phenylketonuria (PKU) was investigated on carnitine metabolism in rats and in patients with PKU. Of phenylacetic acid (PEAA), phenylpyruvic acid and homogentisic acid the PEAA was found to be the most effective in inhibiting carnitine biosynthesis in rats. Following 60 min, a single intraperitoneal dose of PEAA the relative conversion rate, i. e. the hydroxylation, of tracer [Me-(3)H]butyrobetaine to [Me-(3)H]carnitine decreased from 62.2+/-6.00% to 39.4+/-5.11% (means+/-S.E.M., P<0.01) in the liver, in the only organ doing this conversion in rats. The conversion of loading amount of unlabeled butyrobetaine to carnitine was also markedly reduced. The impaired hydroxylation of butyrobetaine was reflected by a reduced free and total carnitine levels in the liver and a reduced total carnitine concentration in the plasma. PEAA decreased the hepatic level of glutamic acid and alpha-ketoglutaric acid (alpha-KG), suggesting a mechanism for the reduced flux through the butyrobetaine hydroxylase enzyme, because alpha-KG is an obligatory co-enzyme. In the plasma and urine of PKU patients on unrestricted diet, markedly decreased total carnitine levels were detected. In the liver of PEAA-treated rats and urine of PKU patients, a novel carnitine derivative, phenacetyl-carnitine was verified by HPLC and gas chromatography-mass spectrometry.     

Regulation of lipid flux between liver and adipose tissue during transient hepatic steatosis in carnitine-depleted rats

Rats with carnitine deficiency due to trimethylhydrazinium propionate (mildronate) administered at 80 mg/100 g body weight per day for 10 days developed liver steatosis only upon fasting. This study aimed to determine whether the transient steatosis resulted from triglyceride accumulation due to the amount of fatty acids preserved through impaired fatty acid oxidation and/or from up-regulation of lipid exchange between liver and adipose tissue. In liver, mildronate decreased the carnitine content by approximately 13-fold and, in fasted rats, lowered the palmitate oxidation rate by 50% in the perfused organ, increased 9-fold the triglyceride content, and doubled the hepatic very low density lipoprotein secretion rate. Concomitantly, triglyceridemia was 13-fold greater than in controls. Hepatic carnitine palmitoyltransferase I activity and palmitate oxidation capacities measured in vitro were increased after treatment. Gene expression of hepatic proteins involved in fatty acid oxidation, triglyceride formation, and lipid uptake were all increased and were associated with increased hepatic free fatty acid content in treated rats. In periepididymal adipose tissue, mildronate markedly increased lipoprotein lipase and hormone-sensitive lipase activities in fed and fasted rats, respectively. On refeeding, carnitine-depleted rats exhibited a rapid decrease in blood triglycerides and free fatty acids, then after approximately 2 h, a marked drop of liver triglycerides and a progressive decrease in liver free fatty acids. Data show that up-regulation of liver activities, peripheral lipolysis, and lipoprotein lipase activity were likely essential factors for excess fat deposit and release alternately occurring in liver and adipose tissue of carnitine-depleted rats during the fed/fasted transition.     

Hydroxylation of gamma-butyrobetaine by rat and ovine tissues.

Ovine tissues were assayed for the capacity to synthesize carnitine from γ-butyrobetaine. Activity in liver, kidney and muscle was 0.25, 0.10 and 0.08 nmoles per mg protein per min, respectively. Heart was devoid of the enzyme. Of the rat tissues that were assayed only liver contained the hydroxylase (0.39 nmoles per mg per min). Although the specific activity of the enzyme was approximately three fold higher in sheep liver than in sheep skeletal muscle, on the basis of total activity, muscle would constitute the major portion of the total hydroxylase activity present in the body. The synthesis of carnitine in ovine skeletal muscle may in part explain the high level of carnitine found in that tissue and emphasizes the existence of species differences in the localization of carnitine synthesis.     

Effect of various levels of supplementation with sodium pivalate on tissue carnitine concentrations and urinary excretion of carnitine in the rat

In previous studies, sodium pivalate has been administered to rats in their drinking water (20 mmoles/L; equivalent to 0.3% of the diet) as a way to lower the concentration of carnitine in tissues and to produce a model of secondary carnitine deficiency. Although this level of supplementation results in a marked decrease in carnitine concentration in a variety of tissues, it does not produce the classical signs of carnitine deficiency (i.e., decreased fatty acid oxidation and ketogenesis). The present study was designed (1) to determine if increasing the level of pivalate supplementation (0.6, 1.0% of the diet) would further reduce the concentrations of total and free carnitine in rat tissues without altering growth or food intake, and (2) to examine the effect of length of feeding (4 vs. 8 weeks) on these variables. Male, Sprague-Dawley rats were randomly assigned to either a control (0.2% sodium bicarbonate) or experimental diet (0.3, 0.6, 1.0% sodium pivalate) for either four or eight weeks. Animals (n = 6/group) were housed in metabolic cages; food and water were provided ad libitum throughout the study. Supplementation with sodium pivalate did not alter water intake or urine output. Ingestion of a diet containing 1.0% pivalic acid decreased food intake (g/day; P < 0.05), final body weight (P < 0.007), and growth rate (P < 0.001) after four weeks. The concentration of total carnitine in plasma, heart, liver, muscle, and kidney was reduced in all experimental groups (P < 0.001), regardless of level of supplementation or length of feeding. The concentration of free carnitine in heart, muscle, and kidney was also reduced (P < 0.001) in rats treated with pivalate for either four or eight weeks. The concentration of free carnitine in liver was reduced in animals supplemented with pivalate for eight weeks (P < 0.05), but no effect was observed in livers from rats treated for four weeks. Excretion of total carnitine and short chain acylcarnitine in urine was increased in pivalate supplemented rats throughout the entire feeding period (P < 0.001). Free carnitine excretion was increased during Weeks 1 and 2 (P < 0.01), but began to decline during Week 3 in experimental groups. During Weeks 6 and 8, free carnitine excretion in pivalate supplemented rats was less than that of control animals (P < 0.01). In summary, no further reduction in tissue carnitine concentration was observed when rats were supplemented with sodium pivalate at levels greater than 0.3% of the diet. Food intake (g/day) and growth were decreased in rats fed a diet containing 1.0% sodium pivalate. These data indicate that maximal lowering of tissue carnitine concentrations is achieved by feeding diets containing 0.3% sodium pivalate or less.     

Carnitine and carnitine orotate affect the expression of the prolactin-releasing peptide gene

Carnitine is involved in fatty acid metabolism in mammals and is widely used as a nutritional supplement; carnitine orotate is a more absorbable form of carnitine. We investigated the effects of carnitine and carnitine orotate on mouse prolactin-releasing peptide (PrRP) mRNA expression. Twenty-four female mice were randomly divided into four groups of six; control mice were orally drenched with physiological saline solution (250 mg/kg body weight) and treatment mice were orally drenched with carnitine (250 mg/kg) or carnitine orotate (250 or 750 mg/kg), once a day, for 20 days from parturition. The carnitine or carnitine orotate was dissolved in saline solution before administration. The hypothalamus, pituitary and ovary were sampled on day 21 after parturition, and PrRP mRNA levels in these tissues were measured by semi-quantitative PCR, with glyceraldehyde 3-phosphate dehydrogenase as a control. Expression of PrRP in mice treated with carnitine and carnitine orotate was significantly increased in the ovary and significantly reduced in the pituitary gland. Compared with the control, hypothalamus PrRP mRNA increased significantly in the carnitine and low-dose carnitine orotate groups and decreased significantly in the high-dose carnitine orotate group. We conclude that carnitine and carnitine orotate regulate expression of PrRP in the pituitary gland and ovaries.     

Tissue carnitine fluxes in vitamin C depleted-repleted guinea pigs

The biosynthesis of carnitine requires vitamin C as a cofactor for two separate hydroxylation steps. The majority of body carnitine (approximately 98%) is located in muscle and less than 0.5% is present in plasma. We examined the physiologic dynamics of plasma free carnitine and muscle total acid-soluble carnitine in vitamin C-depleted guinea pigs repleted with increasing amounts of vitamin C. Animals were fed a vitamin C-deficient diet for 3 weeks at which time symptoms of scurvy were evident. Animals were repleted with increasing doses of vitamin C, from 0.5 to 10.0 mg vitamin C/100 g body weight daily. Muscle total acid-soluble carnitine concentrations tended to correlate directly with plasma vitamin C (r = 0.41, P = 0.087) during the repletion phase of the study. Conversely, plasma free carnitine was inversely related to liver vitamin C (r = −0.54, P = 0.020) and to muscle total acid-soluble carnitine (r = −0.56, P = 0.015). Mean plasma free carnitine values fell 30% over the course of vitamin C repletion (P > 0.05) and mean muscle total acid-soluble carnitine rose by 30% (P > 0.05). These data suggest that elevated plasma free carnitine may indicate a low to marginal vitamin C status.     

Acetyl-L-carnitine supplementation restores decreased tissue carnitine levels and impaired lipid metabolism in aged rats

The effects of long-term carnitine supplementation on age-related changes in tissue carnitine levels and in lipid metabolism were investigated. The total carnitine levels in heart, skeletal muscle, cerebral cortex, and hippocampus were approximately 20% less in aged rats (22 months old) than in young rats (6 months old). On the contrary, plasma carnitine levels were not affected by aging. Supplementation of acetyl-l-carnitine (ALCAR; 100 mg/kg body weight/day for 3 months) significantly increased tissue carnitine levels in aged rats but had little effect on tissue carnitine levels in young rats. Plasma lipoprotein analyses revealed that triacylglycerol levels in VLDL and cholesterol levels in LDL and in HDL were all significantly higher in aged rats than in young rats. ALCAR treatment decreased all lipoprotein fractions and consequently the levels of triacylglycerol and cholesterol. The reduction in plasma cholesterol contents in ALCAR-treated aged rats was attributable mainly to a decrease of cholesteryl esters rather than to a decrease of free cholesterol. Another remarkable effect of ALCAR was that it decreased the cholesterol content and cholesterol-phospholipid ratio in the brain tissues of aged rats. These results indicate that chronic ALCAR supplementation reverses the age-associated changes in lipid metabolism.     

Effect of exogenous carnitine on carnitine homeostasis in the rat

The interaction of exogenous carnitine with whole body carnitine homeostasis was characterized in the rat. Carnitine was administered in pharmacologic doses (0-33.3 mumols/100 g body weight) by bolus, intravenous injection, and plasma, urine, liver, skeletal muscle and heart content of carnitine and acylcarnitines quantitated over a 48 h period. Pre-injection urinary carnitine excretion was circadian as excretion rates were increased 2-fold during the lights-off cycle as compared with the lights-on cycle. Following carnitine administration, there was an increase in urinary total carnitine excretion which accounted for approx. 60% of the administered carnitine at doses above 8.3 mumols/100 g body weight. Urinary acylcarnitine excretion was increased following carnitine administration in a dose-dependent fashion. During the 24 h following administration of 16.7 mumols [14C]carnitine/100 g body weight, urinary carnitine specific activity averaged only 72 +/- 4% of the injection solution specific activity. This dilution of the [14C]carnitine specific activity suggests that endogenous carnitine contributed to the increased net urinary carnitine excretion following carnitine administration. 5 min after administration of 16.7 mumol carnitine/100 g body weight approx. 80% of the injected carnitine was in the extracellular fluid compartment and 5% in the liver. Plasma, liver and soleus total carnitine contents were increased 6 h after administration of 16.7 mumols carnitine/100 g body weight. 6 h post-administration, 37% of the dose was recovered in the urine, 12% remained in the extracellular compartment, 9% was in the liver and 22% was distributed in the skeletal muscle. In liver and plasma, short chain acylcarnitine content was increased 5 min and 6 h post injection as compared with controls. Plasma, liver, skeletal muscle and heart carnitine contents were not different from control levels 48 h after carnitine administration. The results demonstrate that single, bolus administration of carnitine is effective in increasing urinary acylcarnitine elimination. While liver carnitine content is doubled for at least 6 h following carnitine administration, skeletal muscle and heart carnitine pools are only modestly perturbed following a single intravenous carnitine dose. The dilution of [14C]carnitine specific activity in the urine of treated animals suggests that tissue-blood carnitine or acylcarnitine exchange systems contribute to overall carnitine homeostasis following carnitine administration.     

Effect of sports activity on carnitine metabolism: Measurement of free carnitine, γ-butyrobetaine and acylcarnitines by tandem mass spectrometry

The effects of sports activity on carnitine metabolism were studied using mass spectrometry. Serum levels of free carnitine, acylcarnitines (acetylcarnitine, propionylcarnitine, C4-, C5- and C8-acylcarnitine) and γ-butyrobetaine, a carnitine precursor, were determined by tandem mass spectrometry in liquid secondary ion mass ionization mode. The coefficients of variation at three different concentrations were 2.8∼7.9% for γ-butyrobetaine, and 1.2∼6.7% for free carnitine. The recoveries added to serum were 109.1% for γ-butyrobetaine, 89.3% for free carnitine. Sports activity caused increased serum levels of γ-butyrobetaine, acetylcarnitine, C4- and C8-acylcarnitines and decreased serum levels of free carnitine. This method requires a small amount of sample volume (20 μl of serum) and short total instrumental time for the analysis (1 h for preparation, 2 min per sample for mass spectrometric analysis). Therefore, this method can be applied to study carnitine metabolism under various conditions that affect fatty acid oxidation.     

Acid soluble, short chain esterified and free carnitine in the liver, heart, muscle and brain of pre and post hatched chicks

1. The behaviour of total acid soluble, short chain esterified and free carnitine in the liver, heart, muscle and brain of chick embryos between 11th and 21st day of development and of 8 and 180-day-old chicks is described. 2. Total acid soluble carnitine fluctuates around the same levels in the brain, liver and muscle until 18th day of development, whereas it attains a peak on that day in the heart. At hatching compared to 18th day, it suddenly increases three times in the muscle, drops not significantly in the heart and brain, but sharply in the liver (-40%). However the levels are always higher than those of the grown chick in the brain but lower in the other tissues. 3. Free carnitine levels are almost constant in all tissues during the embryonic life; if compared to adult ones, they are very much lower in the liver, heart and muscle, but higher in the brain, even in 8 day-old chick. 4. Short chain esterified, carnitine reaches a maximum on 18th day of egg incubation in the liver, brain and heart; in the muscle it stays on constant levels until this day and then rapidly increases so that at hatching it doubles the values. 5. The short chain esterified to free carnitine percentage ratio peaks in all tissues on 18th day of development, attaining figures which are well above those determined in the grown chick.     

Erythrocyte carnitine: a study of erythrocyte fractions isolated by discontinuous density gradient centrifugation

Erythrocyte fractions of varying density were isolated by discontinuous density gradient centrifugation of washed erythrocytes of five subjects (three adults and two cord bloods). Free and total carnitine concentrations were determined in each gradient fraction to compare the carnitine content of less dense with more dense erythrocytes. Erythrocyte, leukocyte, and reticulocyte counts and hemoglobin were measured on all fractions of each gradient. The density gradient studies showed that the highest proportion of reticulocytes were associated with the least dense gradient fractions of all five subjects. Linear regression analyses revealed significant positive correlations (r = 0.94 to 0.99, P less than 0.02 to P less than 0.001) between the number of reticulocytes per fraction and the total or free carnitine concentrations per fraction for all subjects. No correlation was found between free or total carnitine and hemoglobin, number of erythrocytes, or number of leukocytes per fraction. It appears that erythrocyte carnitine is localized in circulating reticulocytes which have mitochondria and carnitine-dependent fatty acid metabolism.     

The effects of 1-amino-D-proline on the production of carnitine from exogenous protein-bound trimethyllysine by the perfused rat liver

Following receptor-mediated endocytosis of trimethyllysine-labeled asialofetuin and agalacto-orosomucoid by liver parenchymal and nonparenchymal cells, respectively, the glycoproteins are degraded and the methylated lysine residues released. The free intracellular trimethyllysine is then converted, in addition to 2-N-acetyl-6-N-trimethyllysine, to 4-N-trimethylaminobutyrate, carnitine, and acetylcarnitine. In the presence of 1-amino-D-proline, a vitamin B6 antagonist, the total production from protein-bound trimethyllysine of 4-N-trimethylaminobutyrate, the immediate precursor of carnitine, carnitine, and its acetylated derivative was depressed by as much as 60-80% in perfused rat liver. The decreased synthesis of carnitine was accompanied by an accumulation of 3-hydroxy-6-N-trimethyllysine, and intermediate in the carnitine biosynthetic pathway. The extent of 3-hydroxy-6-N-trimethyllysine accumulation, which was not evident in the absence of added 1-amino-D-proline, depended on the dose of 1-amino-D-proline perfused through the liver. In addition, those effects of 1-amino-D-proline were almost completely reversed by inclusion of pyridoxine in the perfusing medium. These results support the suggestion of a requirement for pyridoxal 5'-phosphate in the biosynthesis of carnitine by the liver.     

Relationships between carnitine and coenzyme A esters in tissues of normal and alloxan-diabetic sheep

1. The total acid-soluble carnitine concentrations of four tissues from Merino sheep showed a wide variation not reported for other species. The concentrations were 134, 538, 3510 and 12900nmol/g wet wt. for liver, kidney cortex, heart and skeletal muscle (M. biceps femoris) respectively. 2. The concentration of acetyl-CoA was approximately equal to the concentration of free CoA in all four tissues and the concentration of acid-soluble CoA (free CoA plus acetyl-CoA) decreased in the order liver>kidney cortex>heart>skeletal muscle. 3. The total amount of acid-soluble carnitine in skeletal muscle of lambs was 40% of that in the adult sheep, whereas the concentration of acid-soluble CoA was 2.5 times as much. A similar inverse relationship between carnitine and CoA concentrations was observed when different muscles in the adult sheep were compared. 4. Carnitine was confined to the cytosol in all four tissues examined, whereas CoA was equally distributed between the mitochondria and cytosol in liver, approx. 25% was present in the cytosol in kidney cortex and virtually none in this fraction in heart and skeletal muscle. 5. Carnitine acetyltransferase (EC 2.3.1.7) was confined to the mitochondria in all four tissues and at least 90% of the activity was latent. 6. Acetate thiokinase (EC 6.2.1.1) was predominantly (90%) present in the cytosol in liver, but less than 10% was present in this fraction in heart and skeletal muscle. 7. In alloxan-diabetes, the concentration of acetylcarnitine was increased in all four tissues examined, but the total acid-soluble carnitine concentration was increased sevenfold in the liver and twofold in kidney cortex. 8. The concentration of acetyl-CoA was approximately equal to that of free CoA in the four tissues of the alloxan diabetic sheep, but the concentration of acid-soluble CoA in liver increased approximately twofold in alloxan-diabetes. 9. The relationship between CoA and carnitine and the role of carnitine acetyltransferase in the various tissues is discussed. The quantitative importance of carnitine in ruminant metabolism is also emphasized.     

Developmental changes in the activities of peroxisomal and mitochondrial beta-oxidation in chicken liver

The activities of peroxisomal and mitochondrial beta-oxidation and carnitine acyltransferases changed during the process of development from embryo to adult chicken, and the highest activities of peroxisomal beta-oxidation, palmitoyl-CoA oxidase, and carnitine acetyltransferase were found at the hatching stage of the embryo. The profiles of these alterations were in agreement with those of the contents of triglycerides and free fatty acids in the liver. The highest activities of mitochondrial beta-oxidation and palmitoyl-CoA dehydrogenase were observed at the earlier stages of the embryo; then the activities decreased gradually from embryo to adult chicken. The ratio of activities of carnitine acetyltransferase in peroxisomes and mitochondria (peroxisomes/mitochondria) increased from 0.54 to 0.82 during the development from embryo to adult chicken. The ratio of activities of carnitine palmitoyltransferase decreased from 0.82 to 0.25 during the development. The affinity of fatty acyl-CoA dehydrogenase toward the medium-chain acyl-CoAs (C6 and C8) was high in the embryo and decreased with development, whereas the substrate specificity of fatty acyl-CoA oxidase did not change. The substrate specificity of mitochondrial carnitine acyltransferases did not change with development. The affinity of peroxisomal carnitine acyltransferases toward the long-chain acyl-CoAs (C10 to C16) was high in the embryo, but low in adult chicken.     

Influence of carnitine supplementation on muscle substrate and carnitine metabolism during exercise

We examined 1) the effect of L-carnitine supplementation on free fatty acid (FFA) utilization during exercise and 2) exercise-induced alterations in plasma levels and skeletal muscle exchange of carnitine. Seven moderately trained human male subjects serving as their own controls participated in two bicycle exercise sessions (120 min, 50% of VO2max). The second exercise was preceded by 5 days of oral carnitine supplementation (CS; 5 g daily). Despite a doubling of plasma carnitine levels, with CS, there were no effects on exercise-induced changes in arterial levels and turnover of FFA, the relation between leg FFA inflow and FFA uptake, or the leg exchange of other substrates. Heart rate during exercise after CS decreased 7-8%, but O2 uptake was unchanged. Exercise before CS induced a fall from 33.4 +/- 1.6 to 30.8 +/- 1.0 (SE) mumol/l in free plasma carnitine despite a release (2.5 +/- 0.9 mumol/min) from the leg. Simultaneously, acylated plasma carnitine rose from 5.0 +/- 1.0 to 14.2 +/- 1.4 mumol/l, with no evidence of leg release. Consequently, total plasma carnitine increased. We concluded that in healthy subjects CS does not influence muscle substrate utilization either at rest or during prolonged exercise and that free carnitine released from muscle during exercise is presumably acylated in the liver and released to plasma.     

Distribution of carnitine and acylcarnitine in the hamster epididymis and in epididymal spermatozoa during maturation

The highest levels of carnitine and acylcarnitine were found in the cauda epididymidis, and spermatozoa from the cauda contained greater amounts of total carnitine (free carnitine plus acylcarnitine) than those removed from the corpus or caput epididymidis. Spermatozoa from the distal cauda contained significantly greater amounts of both free and total carnitine than those removed from the proximal cauda epididymidis. The acylcarnitine:carnitine ratio was 1.7 and 0.37 in caput and cauda spermatozoa, respectively and 1.7 and 1.3 in caput and cauda fluid, respectively. It is suggested that the accumulation of carnitine is involved in sperm maturation and that acylcarnitine serves as an energy substrate for epididymal spermatozoa.     

Carnitine metabolism in livers and hearts of 48h-starved rats: the contribution of fat and carbohydrate to acylcarnitine formation

The work investigated the effects of administration of 2-tetradecylglycidate (TDG), an inhibitor of mitochondrial long-chain fatty acid oxidation, alone or in combination with glucose, on concentrations of free and acylated carnitine in livers and hearts of 48 h-starved rats. The only significant effect of TDG in the heart was to decrease [short-chain acylcarnitine]. This demonstrates that in heart, fat oxidation is linked to the formation of short-chain acylcarnitine. Cardiac [short-chain acylcarnitine] was not significantly decreased by TDG if the rats were also administered glucose, suggesting that acyl CoA derived from glucose may be used for short-chain acylcarnitine formation in TDG-treated rats. TDG significantly decreased in [free carnitine]. No changes in [short-chain acylcarnitine] were observed. This indicates that formation of short-chain acylcarnitine in liver is not determined by the rates of fat oxidation. It was calculated that at least 63% of the acyl-groups esterified to carnitine were generated by intramitochondrial beta-oxidation. The effects of glucose and TDG on hepatic concentrations of free and long-chain acylcarnitine were additive, suggesting that extramitochondrial fat oxidation can contribute to acylcarnitine formation in liver.