Plasma Free Carnitine Research Articles

A study of the distribution of total, free, short-chain acyl and long-Chain acyl carnitine in whole blood and plasma of arabian sand gazelles (Gazella subgutturosa marica)

This study was conducted to investigate the presence of total carnitine, free carnitine, short-chain acyl carnitine, long-chain acyl carnitine, acyl carnitine levels and the ratio of acyl carnitine to free carnitine in erythrocytes, whole blood and plasma of Arabian sand gazelles (Gazella subgutturosa marica). Results showed erythrocyte levels of carnitine significantly higher compared to either whole blood or plasma of Arabian sand gazelles (p<0.001). Moreover, the relative distribution of acyl carnitine was higher in erythrocytes compared to most species (p<0.001), however, it was similar to the Arabian camel (p<0.001). The ratio of acyl carnitine to free carnitine was significantly higher in erythrocytes, whole blood and plasma compared to most species (p<0.001). The observed discrepancies in carnitine levels among species and different tissues is well known, however, the reasons for these discrepancies are obscure. The higher carnitine content and a higher acyl carnitine to free carnitine ratio in blood of Arabian sand gazelles may suggest that these animals possess an adaptive mechanism which may be found in other desert animal species. This is the first report describing deer carnitine distributions.

THE IDENTIFICATION OF A NOVEL G4.5 MUTATION IN BARTH SYNDROME• 96

Barth syndrome is an X-linked recessive cardioskeletal myopathy associated with neutropenia, growth retardation, abnormal mitochondria (abnormal cristae), and elevated levels of urinary 3-methylglutaconic acid and 2-ethylhydracrylic acid. Lipid myopathy, decreased plasma free carnitine, lowered cytochrome concentrations, and granulocytopenia are also characteristic of this disease. The gene for Barth syndrome, which was previously mapped to Xq28, was recently identified as G4.5. This 6,234 bp gene consists of 11 exons, three of which (exons 5, 6, and 7), are differentially spliced resulting in five protein products. These proteins, classified as tafazzins, show no homolgy to any previously identified proteins, and thus, are unique. Four mutations in G4.5 have previously been described in patients with Barth syndrome. We report a new mutation in G4.5 in one family having three males affected with Barth syndrome.

Cerebrospinal fluid carnitine levels in patients with Parkinson's disease

We assessed free carnitine (FC) and acyl-carnitine esters (AC) in both CSF and plasma from 29 patients with diagnostic criteria for PD, and from 29 healthy matched-controls. FC and AC levels in both CSF and plasma did not differ significantly between PD patients and controls, they were not influenced significantly by anti-parkinsonian drugs, and did not correlate with age at onset, duration and severity of PD. These results suggest that CSF carnitine levels are apparently unrelated with the risk for PD. © 1997 Elsevier Science B.V. All rights reserved.

Plasma Carnitine Kinetics during Orthotopic Liver Transplantation

Background: Carnitine is synthesized mainly in the liver and plays an essential role in the transport of fatty acids in liver mitochondria for subsequent oxidation and energy production. Methods: The plasma concentrations of free carnitine, acylcarnitine, total ketone bodies, lactate, pyruvate, and hepatocyte growth factor (HGF) were measured during liver transplantation. Results: The plasma free carnitine and acylcarnitine concentrations and the lactate to pyruvate ratio in patients with compromised grafts (group A) were significantly higher than those in patients with well-functioning grafts (group B) after reperfusion. The acylcarnitine concentration in group B decreased after incision, but it remained at a high level in group A. Significant correlations were found between the concentrations of HGF and free and acylcarnitine after reperfusion. Conclusion. The accelerated flux of carnitine in the graft may be associated with deterioration of energy metabolism in the graft. An increased acylcarnitine concentration may reflect impaired liver regeneration.

Cytotoxic Aldehydes as Possible Markers for Childhood Cancer

Concentrations of 22 known aldehydes (byproducts of lipid peroxidation), 5 acyloins, free and total carnitine and acylcarnitines were measured in plasma and urine obtained from pediatric patients with various forms of cancer before any treatment, and following treatment with doxorubicin or daunorubicin. Aldehydes, before the initiation of chemotherapy, were significantly elevated in cancer patients compared to controls. Aldehydes such as hexanal, heptanal, and malondialdehyde were strikingly higher in samples from cancer patients, while trans 4-cis-4-decenal was the prominent aldehyde in the blood of controls. In addition, in each form of cancer the pattern of aldehydes appeared to be unique when compared to controls, or to others forms of cancer. In cancer patients receiving chemotherapy there was a general trend toward a reduction 24 h after both the first and after the fifth doxorubicin dose. These changes however were not significant statistically due to large inter-patient variation. Free and total plasma carnitine levels remained in the normal range, and there were no abnormal acylcarnitines detected in urine. Possible hypotheses to explain the elevations in aldehydes, and the reasons for the changed aldehyde profiles in different forms of cancer are discussed.

Vitamin C depletion is associated with alterations in blood histamine and plasma free carnitine in adults.

The purpose of this study was to determine whether carnitine metabolism or histamine degradation would be useful parameters for investigating the optimal requirement for vitamin C. Twenty-two non-scorbutic subjects with subnormal vitamin C status (plasma vitamin C < 28 mumol/L) were placed on a metabolic diet low in vitamin C for 3 weeks and repleted with graded doses of vitamin C: 10, 30 and 60 mg vitamin C daily (group 1) or 10,125 and 250 mg vitamin C daily (group 2) for weeks 1, 2 and 3, respectively. Fasting blood samples were collected weekly and analyzed for plasma vitamin C, plasma free carnitine and blood histamine. Group 1 subjects remained in a subnormal vitamin C state throughout the 3-week study, and blood histamine and plasma free carnitine were not impacted by the experimental treatment. Plasma vitamin C in group 2 subjects rose significantly during the study, and these subjects finished the study with an ample vitamin C status indicative of vitamin C intakes above the recommended dietary allowance. Both blood histamine and plasma free carnitine were inversely related to vitamin C status in group 2 subjects. These data indicate that blood histamine and plasma free carnitine are altered in individuals with subnormal, non-scorbutic vitamin C status and provide evidence that metabolic changes independent of collagen metabolism occur prior to the manifestation of scurvy. Thus utilizing scurvy as an end-point to determine vitamin C requirements may not provide adequate vitamin C to promote optimal health and well-being.

Pivalate Affects Carnitine Status but Causes No Severe Metabolic Changes in Rat Liver

To investigate the carnitine deficiency induced by pivalate, rats had free access to drinking water with or without pivalate. Consumption of 20 mmol/L pivalate for 1 wk decreased the levels of both free and total carnitine in plasma to ∼20% of levels before treatment. After 4 wk, the concentrations of free carnitine in the liver, heart and muscle of pivalate-treated rats were ∼60–80% of the control, and in the kidney, 26% of the control. Fractional excretion of free carnitine (FEFC) in pivalate-treated rats was measured; however, the treatment for 3 or 8 d did not affect the values relative to those obtained before treatment. Treatment with pivalate for 4 wk did not affect plasma concentrations of glucose, ammonia and free fatty acids (FFA) in the rats; however, the concentration of 3-hydroxybutyrate (3-OHB) was higher, and the FFA/3-OHB ratio was lower than those of controls. In a liver perfusion study, ketogenesis from oleate and gluconeogenesis from lactate and pyruvate in rats treated with pivalate for 4 wk were not different from controls. These results suggest that administration of pivalate did not induce the excessive excretion of free carnitine in urine, and secondary carnitine deficiency induced by intake of 20 mmol/L pivalate for 4 wk did not cause severe metabolic changes in rat liver.

Free and esterified carnitine in continuous ambulatory peritoneal dialysis patients

Free, acetyl-, medium- and long-chain acylcarnitine and total plasma carnitine concentrations were measured in eight continuous ambulatory peritoneal dialysis (CAPD) patients and eight age- and sex-matched healthy controls. Daily loss of carnitine was also quantified in both groups, by analysis of urine and dialysis fluid. Plasma total carnitine concentration in CAPD patients was not significantly different from controls (42.8 +/- 1.6 and 43.1 +/- 2.3 mumol/liter, respectively). However, the plasma free carnitine concentration of CAPD patients was significantly lower than that of controls (28.5 +/- 1.4 and 36.2 +/- 2.5 mumol/liter, respectively; P < 0.05). No difference in the daily loss of total carnitine was found between CAPD patients and controls (269.7 +/- 30.0 and 240.5 +/-33.0 mumol/liter, respectively), but the daily loss of free carnitine was significantly greater in CAPD patients (175.8 +/- 17.3 and 105.8 +/- 16.4 mumol/liter, respectively; P < 0.05). The ratio of total acylcarnitine (acetyl-, medium- and long-chain acylcarnitine) to free carnitine was significantly greater in plasma of CAPD patients than in controls (P < 0.01) and was lower in daily fluid losses (P < 0.001). These ratio differences suggests that an alteration in acyl group metabolism is occurring in CAPD patients. This may be attributable to an accumulation of medium- and long-chain acylcarnitine in liver of CAPD patients which would be exchanged for plasma free carnitine and/or to a differential loss of free and acylcarnitine across the peritoneal cavity.

HPLC determination of carnitine and acylcarnitines in human plasma by means of fluorescence labeling using 2-(4-hydrazinocarbonylphenyl)-4,5-diphenylimidazole.

Carnitine and acylcarnitines are important substances involved in the oxidation and metabolism of fatty acids. An HPLC method is presented for the quantitative analysis of these compounds. The method is based on the detection of fluorescent derivatives of carnitine and short- and medium-chain acylcarnitines labeled with 2-(4-hydrazinocarbonylphenyl)-4,5-diphenylimidazole (HCPI). The labeling of carnitine and acylcarnitines with HCPI was performed at room temperature for 1 h using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide as a condensing reagent. The analytes prepared using cation-exchange cartridges were separated on an octadecylsilyl silica gel (ODS) column by a gradient elution system of acetonitrile/Tris-HCI buffer and determined using a synthetic internal standard. Fluorescence detection was performed at 475 nm with excitation at 340 nm. The detection limits for carnitine, acetyl-, propionyl-, hexanoyl- and octanoylcarnitine ranged from 0.24 to 1.97 nmol per ml human plasma, at a signal-to-noise ratio of 3. The within-day and between-day precision of the assay for carnitine and acylcarnitines in plasma samples had relative standard deviations (RSDs) lower than 10.3%. The concentration of free carnitine, acylcarnitines and total carnitine in human plasma could be successfully determined by the proposed method.

Effects of parenteral L-carnitine supplementation on fat metabolism and nutrition in premature neonates

The effects of parenteral L-carnitine supplementation on fat metabolism, nutrient intake, and plasma and erythrocyte carnitine concentrations were studied in 43 very low birth weight infants. Infants were randomly assigned to control or carnitine-supplemented (50 μmol/kg per day) groups within two weight categories: group 1, 750 to 1000 gm, and group 2, 1001 to 1500 gm. Plasma total, free, and acyl carnitine levels, erythrocyte carnitine levels, serum β-hydroxybutyrate and triglyceride levels, and total fat intake were monitored weekly until 50% of total caloric intake was met enterally. Neonates receiving carnitine had higher plasma carnitine levels than control groups (total carnitine: group 1, 75.2 ± 22.9 vs 9.6 ± 2.7 mmol/ml; group 2, 61.6 ± 31.2 vs 13.0 ± 9.2 nmol/ml). Levels of β-OH-butyrate decreased from baseline in control neonates (group 1, 0.12 ± 0.06 to 0.03 ± 0.02 mmol/L; group 2, 0.11 ± 0.03 to 0.05 ± 0.02 mmol/L); they remained unchanged in supplemented groups. Thus ketogenesis appeared less impaired in infants receiving supplements. Supplemented group 2 tolerated more fat than control group 2; triglyceride levels remained acceptable in all groups. Carnitine group 2 had greater weight gain than control group 2 during the first 2 weeks of life. We conclude that very low birth weight infants requiring prolonged parenteral nutrition have carnitine deficiency with impaired ketogenesis. Parenteral administration of carnitine appears to alleviate this metabolic disturbance. (J P EDIATR 1995;126:287-92)

227 PLASMA FREE CARNITINE LEVELS IN CHILDREN WITH MALNUTRITION

In this study plasma free carnitine and albumin levels were measured in children with protein energy malnutrition (PEM). A total of 71 children with malnutrition were studied. The control group consisted of 20 healthy children. Mean plasma carnitin leve1 was 78.4±1.94 nmol/ml in the control group. Marasmus and kwashiorkor patients displayed lower plasma free carnitine values which were found statistically significant when compared to those of the control ( P 5.46). These was no correlation between serum albumin and free carnitine levels in cases with kwashiorkor. The reason why no correlat ion between serum a 1bumin and free carnitine levels was observed in the cases investigated in this work has been attributed to varying dis f unc t ion of the control mechanism in cases with kwashiorkor. We concluded that further inves tigations of carnitine concentration at tissue level are essential in PEM for a more effective manegement of carnitine deficiency by oral means.

Acylcarnitine profile in tissues and body fluids of biotin-deficient rats with and without L-carnitine supplementation.

Since biotin-deficient (BD) rats are a good animal model for human multiple carboxylase deficiency and have low plasma free carnitine levels, short-chain acylcarnitine profiles in biotin-deficient rats with L-carnitine supplementation (BDC rats) and BD rats were investigated by fast-atom bombardment and tandem mass spectrometry and gas chromatography/mass spectrometry. By the latter method, 3-hydroxyisovalerylcarnitine was identified in BD rats, and showed the greatest accumulation among short-chain acylcarnitines in tissues of BD rats, while the tissue levels of propionic acid were more markedly elevated than those of 3-hydroxyisovaleric acid. The tissue levels of 3-hydroxyisovaleryl-carnitine were significantly lower and those of propionyl-carnitine were somewhat higher in BDC rats than in BD rats, while the tissue levels of propionic acid and 3-hydroxyisovaleric acid in BDC rats were lower than those in BD rats. These changes were more apparent in kidney than in other tissues. The amounts of urinary excretion of acylcarnitines were markedly larger, and those of 3-hydroxyisovaleric acid were somewhat smaller in BDC rats than in BD rats, while those of propionic acid were very low in BD and BDC rats as compared with those of 3-hydroxyisovaleric acid. It seems that the relationship between the concentrations of 3-hydroxyisovalerylcarnitine and those of propionylcarnitine reflects the unique metabolism of the related metabolites in tissues, especially in kidney, which may be influenced by their urinary excretion and the availability of free carnitine. These data in biotin deficiency suggest that carnitine supplementation is possibly beneficial for patients with holocarboxylase synthetase deficiency who respond incompletely to biotin therapy.

Renal adaptation to dietary carnitine in humans

Carnitine homeostasis in humans is maintained by dietary carnitine intake, a modest rate of endogenous carnitine synthesis, and efficient conservation of carnitine by the kidney. To assess the effect of dietary carnitine on the efficiency of carnitine reabsorption in humans, rates of carnitine excretion and reabsorption, indexed to the glomerular filtration rate, were determined over a range of plasma free and total carnitine concentrations in 12 strict vegetarians before and after dietary carnitine supplementation (0.248 mmol/d). This amount of dietary carnitine supplementation did not significantly increase plasma carnitine concentration and did not alter the glomerular filtration rate. At normal physiological plasma carnitine concentrations, the rate of carnitine excretion was increased and the rate of carnitine reabsorption was decreased by carnitine supplementation. We conclude that the kidney adapts to carnitine intake by reducing the efficiency of carnitine reabsorption.

Impaired fatty acid oxidation in children on valproic acid and the effect of L‐carnitine

Fatty acid oxidation was studied in 12 patients (aged 3 to 19 years) receiving valproic acid (VPA), predominantly as monotherapy, before and after 1 month of L-carnitine supplementation (50 mg/kg/day po) in order to determine whether L-carnitine plays a role in preventing the hepatotoxic effects of this drug. Five of these patients were also studied prior to VPA treatment. Only one patient taking VPA had an abnormally low plasma free carnitine. Acyl-/free carnitine ratios were elevated in five patients on VPA and normalized after L-carnitine supplementation. Mean plasma concentrations of free fatty acids, beta-OH-butyrate, and cumulative excretion of 13CO2 after administration of 1-13C-octanoic acid were not changed by VPA or L-carnitine treatment. Urinary dicarboxylic acids, acylglycines, and octanoylcarnitine were elevated during VPA therapy and unaltered by L-carnitine. These results suggest that, in patients at low risk for VPA-induced hepatotoxicity (patients aged > 2 years and taking VPA as monotherapy), VPA causes metabolic abnormalities resembling those found in inborn errors of mitochondrial beta-oxidation which are not corrected by L-carnitine.

Relationship of Carnitine and Carnitine Precursors Lysine, ɛ-N-Trimethyllysine, and γ-Butyrobetaine in Drug-Induced Carnitine Depletion

Plasma concentrations and rates of urinary excretion of carnitine and some of its precursors were studied in three groups of children receiving drugs known to cause carnitine depletion. Patients in group A received pivampicillin and a molar equivalent of carnitine for 7 d. Patients in group B received pivampicillin with a 5.8-fold molar excess of carnitine for 1 wk. Patients in group C were treated chronically with valproic acid and received a molar equivalent (to valproic acid) of carnitine for 14 d. Patients in group A had markedly increased (16-fold) urinary carnitine ester excretion concomitant with diminished urinary free carnitine and gamma-butyrobetaine output and lower plasma free carnitine concentration. Supplementation with one molar equivalent of carnitine (to pivampicillin) was ineffective in preventing the reduction of plasma carnitine concentration observed with pivampicillin treatment alone. For group B patients, administration of excess carnitine resulted in a further increase (35-fold) of urinary carnitine ester output with no decrease of plasma carnitine concentration, urinary gamma-butyrobetaine, or free carnitine excretion. For patients in group C, the initially low plasma free and total carnitine concentrations and urinary output of carnitine and carnitine esters markedly increased with carnitine supplementation, but urinary excretion of gamma-butyrobetaine remained unchanged. The plasma concentrations and urinary output of L-lysine and epsilon-N-trimethyllysine remained unchanged within each group before and after treatment. A positive linear correlation was found between urinary epsilon-N-trimethyllysine and 3-methylhistidine output, indicating that the rate of epsilon-N-trimethyllysine excretion correlates with the amount of 3-methylhistidine liberated by protein turnover.(ABSTRACT TRUNCATED AT 250 WORDS)

Muscle carnitine repletion by long-term carnitine supplementation in nephropathic cystinosis.

The renal tubular Fanconi syndrome of children with nephropathic cystinosis causes plasma and muscle carnitine depletion. L-Carnitine replacement therapy for up to 18 mo has previously been shown to normalize plasma but not muscle carnitine levels. We treated six cystinosis patients, aged 1 to 4 y, with a mean dosage of 92 mg L-carnitine/kg/d given every 6 h for an average of 62 mo. Despite fractional excretions of free carnitine ranging from 55 to 108%, plasma-free and total carnitine concentrations were maintained at or above normal levels. At the end of the carnitine replacement period, the six children had muscle-free carnitine values ranging from 16.0 to 28.0 nmol/mg noncollagen protein compared with values of 3.0 to 11.4 for cystinosis children not supplemented with carnitine [normal, 22.7 +/- 5.0 (SD) nmol/mg protein]. Total muscle carnitine values were also normalized by L-carnitine replacement. The monthly increase in total body creatinine production, a measure of muscle mass, was higher (p = 0.036) in children with normal plasma free carnitine concentrations (3.4 +/- 0.9 mg/d) than in children with low plasma free carnitine (2.3 +/- 0.7 mg/d). No serious side effects, such as severe diarrhea, were observed. We conclude that oral L-carnitine replacement can normalize muscle carnitine content in children with cystinosis.

Quantification of Free Carnitine, Individual Short- and Medium-Chain Acylcarnitines, and Total Carnitine in Plasma by High-Performance Liquid Chromatography

This paper describes a method for the quantitative determination of free carnitine, acetylcarnitine, propionylcarnitine, hexanoylcarnitine, octanoylcarnitine, and total carnitine in plasma. Carnitine and acylcarnitines were extracted from 100 μl of plasma with acetonitrile/methanol and isolated using 0.5-ml columns of silica gel. Samples were then derivatized with 4′-broniophenacyl trifluoromethanesulfonate and quantified by high-performance liquid chromatography with detection at 260 nm. Carnitine and acylcarnitines were quantified in normal human plasma and the plasma of patients diagnosed with methylmalonic aciduria, propionic acidemia, and medium-chain acyl-CoA dehydrogenase deficiency.

Renal Handling of Carnitine in Secondary Carnitine Deficiency Disorders

Reduced plasma and tissue concentrations of carnitine, a cofactor required for fatty acid oxidation, are present in patients with inherited disorders of mitochondrial acyl-CoA oxidation that are associated with accumulations of acylcarnitines. To determine whether the secondary carnitine deficiency in these patients is due to excessive urinary loss of acylcarnitines, the development of carnitine deficiency was examined in patients with four different acyl-CoA oxidation disorders, including medium-chain and long-chain fatty acyl-CoA dehydrogenase deficiencies, isovaleric acidemia, and propionic acidemia. After a 3-mo period of treatment with oral carnitine to raise plasma total carnitine concentrations to or above normal, patients were started on a carnitine-free diet and the changes in plasma total and free carnitine levels and urinary total and free carnitine excretion were followed for 5 d. Patients with all four disorders showed a return of plasma carnitine levels and urinary carnitine excretion to baseline within 2 to 4 d. The rapidity of these changes could not be explained solely by excessive acylcarnitine wasting. Continued excretion of free carnitine in all patients indicated the additional presence of an impairment in renal transport of free carnitine. Consistent with this interpretation, estimates of renal thresholds for free carnitine gave values that were less than that for a control child in all four disorders and ranged as low as one half those reported in normal individuals. These results suggest that secondary carnitine deficiency in the acyl-CoA oxidation disorders is due to indirect as well as direct effects of accumulated acylcarnitines.(ABSTRACT TRUNCATED AT 250 WORDS)

Myocardial carnitine metabolism in congestive heart failure induced by incessant tachycardia.

Persistent tachycardia induces congestive heart failure (CHF), but the mechanism(s) of progressive ventricular dysfunction is (are) unclear. This study was designed to define possible metabolic causes of myocardial dysfunction in rapid ventricular pacing induced CHF. Twelve adult mongrel dogs were paced to 250 beats/min for 19 days. Plasma carnitine, norepinephrine and renin were measured at 0, 1, 2, and 3 weeks. Myocardial high energy phosphates, carnitine, glycogen, glucose, non-collagenous protein and collagen were measured at 19 days. Cardiac output, arterial pressure and pulmonary wedge pressure, measured at baseline and with CHF, showed a decrease in cardiac output and increase in pulmonary wedge pressure. Neurohumoral activation was evident by progressively increasing plasma norepinephrine and renin activity and depletion of myocardial norepinephrine. Plasma free carnitine rose significantly from 12.6 +/- 2.0 control to 28.3 +/- 3.8 nmol/ml at 19 days (p < 0.001), whereas myocardial total carnitine was lower in paced than in control dogs (6.0 +/- 1.9 vs. 14.1 +/- 3.5 nmol/mg non-collagenous protein, p < 0.001). Myocardial ATP ATP and ADP were unchanged, while AMP decreased 22%, and creatine phosphate decreased 30% compared to control animals. Myocardial glucose was normal but glycogen was decreased 54% (p < 0.005). The low myocardial carnitine and elevated plasma carnitine in pacing induced CHF suggests altered carnitine transport or membrane integrity.

Carnitine-acylcarnitine translocase deficiency with severe hypoglycemia and auriculo ventricular block. Translocase assay in permeabilized fibroblasts.

Deficiency of the enzymes of mitochondrial fatty acid oxidation and related carnitine dependent steps have been shown to be one of the causes of the fasting-induced hypoketotic hypoglycemia. We describe here carnitine-acylcarnitine translocase deficiency in a neonate who died eight days after birth. The proband showed severe fasting-induced hypoketotic hypoglycemia, high plasma creatine kinase, heartbeat disorder, hypothermia, and hyperammonemia. The plasma-free carnitine on day three was only 3 microM, and 92% of the total carnitine (37 microM) was present as acylcarnitine. Treatments with intravenous glucose, carnitine, and medium-chain triglycerides had been tried without improvements. Measurements in fibroblasts confirmed deficient oxidation of palmitate and showed normal activities of the carnitine palmitoyltransferases I and II and of the three acyl-CoA dehydrogenases. A total deficiency of the carnitine-acyl-carnitine translocase was found in fibroblasts using the carnitine acetylation assay (1986. Biochem. J. 236:143-148). This assay has been further simplified by seeking conditions permitting application to permeabilized fibroblasts and lymphocytes.