L-carnitine Concentrations Research Articles

L-CARNITINE ENHANCING ROLES ON BUFFALO SEMEN FREEZABILITY, ULTRA STRUCTURE AND FERTILIZING POTENTIALS

Recently, male impact has been recognized as a momentous cause of infertility. Male infertility isn’t an entity but it evokes a variety of mechanisms and explanations to understand it. In between, a biochemical semen additive that improves or overcome certain semen situations so can enhance or treat semen fertility and infertility, respectively. L-carnitine is one of these biochemical semen additives. Till now, the exact effects of L-carnitine on buffalo semen processing outcomes haven’t been discovered. The current study aimed to clarify L-carnitine roles during buffalo semen cryopreservation. Semen samples were obtained from six fertile buffalo bulls (aged 3 to 5 years). Weekly, two consecutive ejaculates were collected from each bull for successive six weeks using artificial vagina. The ejaculates were pooled to eliminate samples variability. Semen samples were extended with Tris-based extender supplemented with different concentrations of L-carnitine (0.01, 0.05 and 0.1 mg/ml) Vs. Tris-based extender only (control). Then they were processed to cryopreservation and thawing to assess different semen characteristics. It had been found that L-carnitine (0.05mg/ml) significantly (p < 0.05) improved post thawing motility, viability index, acrosomal integrity, in vitro fertilization, blastocyst and conception rates of treated buffalo semen (57.00±2.55%, 126.5±8.33, 14.40 ± 1.43%, 56.45%, 15.79% and 64.71%, respectively), compared with control (42.00±2.56%, 87.00±10.43, 23.80±1.86%, 36.53%, 2.13% and 42.86%, respectively). Conclusion: L-carnitine supplementation to buffalo semen extender significantly enhanced its characteristics and protecting its plasma membrane and mitochondrial functional integrity. Moreover, the preceding results focusing more light on the potential roles of L-carnitine in regulating male fertility and infertility.

L-carnitine Mediated Reduction in Oxidative Stress and Alteration in Transcript Level of Antioxidant Enzymes in Sheep Embryos Produced In Vitro.

The objective of this study was to find out the effect of L-carnitine on oocyte maturation and subsequent embryo development, with L-carnitine-mediated alteration if any in transcript level of antioxidant enzymes (GPx, Cu/Zn-SOD (SOD1) and Mn-SOD (SOD2) in oocytes and developing sheep embryos produced in vitro. Different concentrations of L-carnitine (0 mm, 2.5 mm, 5 mm, 7.5 mm and 10 mm) were used in maturation medium. Oocytes matured with 10 mm L-carnitine showed significantly (p < 0.05) higher cleavage (66.80% vs 39.66, 41.76, 44.64, 64.31%), morula (48.50% vs 20.88, 26.01, 26.99, 44.72%) and blastocyst (33.22% vs 7.66, 9.19, 10.71, 28.57%) percentage as compared to lower concentrations (0 mm, 2.5 mm, 5 mm and 7.5 mm). Cleavage percentage between 10 mm and 7.5 mm L-carnitine were not significantly different. Maturation rate was not influenced by supplementation of any experimental concentration of L-carnitine. There was a significant (p < 0.05) decrease in intracellular ROS and increase in intracellular GSH in 10 mm L-carnitine-treated oocytes and embryos than control group. Antioxidant effect of L-carnitine was proved by culturing oocytes and embryos with H2O2 in the presence of L-carnitine which could be able to protect oocytes and embryos from H2O2-induced oxidative damage. L-carnitine supplementation significantly (p < 0.05) upregulated the expression of GPx and downregulated the expression of SOD2 genes, whereas the expression pattern of SOD1 and GAPDH (housekeeping gene) genes was unaffected in oocytes and embryos. It was concluded from the study that L-carnitine supplementation during in vitro maturation reduces oxidative stress-induced embryo toxicity by decreasing intracellular ROS and increasing intracellular GSH that in turn improved developmental potential of oocytes and embryos and alters transcript level of antioxidant enzymes.

Impact of Vigorous exercise on serum levels of L-Carnitine in prisoners in Colombia

Objective To assess the effect of a program of vigorous physical exercises on the serum concentration of free and total L-carnitine, in male inmates at a prison in Boyacá, Colombia. Methods Pre-post intervention population-based study. 44 male prisoners with overweight and/or obesity, from a jail in Boyacá, Colombia were randomly assigned into two groups: an intervention group and a control group. The intervention consisted in participating in a vigorous exercise program over twelve weeks. Anthropometric measures and levels of free and total L-carnitine were every four weeks. Results There were significant increases in serum levels of free and total L-carnitine in the intervention group compared to the control group. Concurrently, in this group there was a reduction in body mass index (BMI), while in the control group there were no changes. Conclusion In overweight and/or obese patients, the routine practice of vigorous exercise plus caloric restriction offers significant benefits in reducing body fat volumes through the mechanisms of energetic consumption of long chain fatty acids.

Correlation between serum L-carnitine concentration and neutrophil engraftment in patients treated with cord blood transplantation

In cord blood transplantation (CBT), the amount of time elapsing until hematological engraftment has effects on the transplantation results. Carnitine deficiency has been reported to cause erythropoietin refractory anemia in chronic hemodialysis patients and thrombocytopenia or leukopenia of cirrhosis, and carnitine supplementation can improve hematopoiesis in patients with hepatic or renal failure. Patients who receive CBT may suffer from carnitine deficiency, but no studies have investigated the carnitine status of such patients. Herein, we determined the concentration of free carnitine (FC) and investigated the correlation between FC and engraftment in patients who received CBT. Twenty-three patients who received CBT at our hospital during the period from April 2013 to January 2015 were enrolled in this study. One patient was excluded because of graft failure, such that 22 patients were ultimately evaluable. FC concentrations of the patients were sequentially monitored at 4 time points (before conditioning therapy, day 0, day 7, and day 14), basic laboratory data were collected, and their correlations with engraftment were analyzed. FC concentrations of the patients were generally low (before conditioning therapy: 33.1, day 0: 43.2, day 7: 38.3, and day 14: 37.8 μmol/l). Significant inverse correlations were observed between FC concentrations and the number of days required for neutrophil engraftment on day 0 and day 14 (before conditioning therapy: P=0.15, r=-0.33, day 0: P=0.04, r=-0.43, day 7: P=0.30, r=-0.23, and day 14: P=0.01, r=-0.55). These results suggest carnitine to be an important nutrient that promotes hematopoietic recovery after CBT.

Effects of L-carnitine against H2O2-induced oxidative stress in grass carp ovary cells (Ctenopharyngodon idellus).

This study was designed in vitro to investigate the effects of L-carnitine against H2O2-induced oxidative stress in a grass carp (Ctenopharyngodon idellus) ovary cell line (GCO). GCO cells were pre-treated with different concentrations of L-carnitine, followed by incubation with 2.5mM H2O2 for 1h to induce oxidative damage. The results indicated that adding L-carnitine at concentrations of 0.01-1mM into the medium for 12h significantly increased cell viability. Pre-treatment with L-carnitine at concentrations of 0.1-5mM for 12h significantly inhibited 2.5mM H2O2-induced cell viability loss. The significant decreases in the level of reactive oxygen species and cell apoptosis were observed in 0.5mM L-carnitine group compared to the H2O2 group. Malondialdehyde values of all of the L-carnitine groups were significantly lower than those of the H2O2 group, while total glutathione levels of all of the L-carnitine groups were significantly higher than of the H2O2 group. The activity of antioxidant enzymes, such as total superoxide dismutase (0.1 and 0.5mM L-carnitine), catalase (0.5mM L-carnitine) and γ-glutamyl cysteine synthetase (0.5 and 1mM L-carnitine), was significantly increased. In addition, pre-treatment of L-carnitine in GCO cells exposed to 2.5mM H2O2 significantly increased the mRNA expression of copper, zinc superoxide dismutase, catalase (0.5mM L-carnitine), glutamate cysteine ligase catalytic subunit (0.1-1mM) and glutathione peroxidase (0.1mM L-carnitine). In conclusion, L-carnitine promotes GCO cell growth and improves antioxidant function, it plays a protective role against oxidative stress induced by H2O2 in GCO cells, and the appropriate supplemental amount of L-carnitine is 0.1-1mM.

163 EFFECTS OF SERUM AND L-CARNITINE ON DEVELOPMENT AND CRYOTOLERANCE OF BOVINE EMBRYOS PRODUCED IN VITRO

Cryotolerance of bovine embryos produced in vitro (PIV) can be improved by l-carnitine. The objective of the present study was to determine whether the optimal concentration of l-carnitine is dependent on serum. Bovine embryos were produced in vitro with abattoir-derived oocytes. After fertilization (Day 0), oocytes (n = 2768) were randomly assigned in a 2 × 4 factorial design to culture in SOF-BE1 medium supplemented with or without 5% fetal bovine serum and l-carnitine at concentrations of 0.0, 0.75, 1.5, and 3.03 mM at 38.5°C in a humidified atmosphere of 5% O2, 5% CO2, and 90% N2. The proportion of oocytes that cleaved was assessed on Day 3, and the proportion of oocytes that developed to the blastocyst and advanced blastocyst (expanded, hatching, and hatched) stages was determined on Day 7. Blastocysts and expanded blastocysts (n = 466) were harvested on Day 7 and subjected to controlled-rate freezing following equilibration in 1.5 M ethylene glycol. After thaw, embryos were cultured for 72 h in SOF-BE1 supplemented with 10% (v/v) fetal bovine serum and 50 mM dithiothreitol at 38.5°C in a humidified atmosphere of 5% O2, 5% CO2, and 90% N2. Post-thaw re-expansion and hatching rates were determined at 24, 48, and 72 h. The experiment was replicated 9 times, and data were analysed by logistic regression. There was no interaction between serum and l-carnitine, at any of the concentrations tested, on embryo development or cryotolerance. Cleavage rates were not affected by serum or l-carnitine. Addition of serum during culture increased (P &lt; 0.05) development to the blastocyst (19.7 ± 1.1% v. 25.3 ± 1.4%) and advanced blastocyst (9.1 ± 0.8% v. 12.4 ± 1.2%) stages. While l-carnitine did not affect blastocyst development, advanced blastocyst development was reduced (P &lt; 0.05) for l-carnitine at 3.03 mM (0 mM: 10.9 ± 1.2%, 0.75 mM: 12.2 ± 1.4%, 1.5 mM: 13.5 ± 1.5%, 3.03 mM: 7.0 ± 1.0%). Serum reduced (P &lt; 0.01) re-expansion (78.1 ± 3.4% v. 65.5 ± 3.1%, 81.0 ± 3.0% v. 68.4 ± 2.7%, 78.4 ± 3.4% v. 65.8 ± 3.1%, for 24, 48, and 72 h, respectively) and hatching (52.0 ± 4.0% v. 39.8 ± 3.6%, 61.2 ± 4.1% v. 45.4 ± 3.8%, 61.2 ± 4.1% v. 45.4 ± 3.8%, for 24, 48, and 72 h, respectively) rates at all time points. In contrast, treatment of embryos with l-carnitine during culture increased (P &lt; 0.05) post-thaw re-expansion rates at 24 and 48 h, regardless of concentration (Table 1). In conclusion, post-thaw viability of bovine embryos PIV can be improved by the addition of l-carnitine during culture. Moreover, the beneficial effects of l-carnitine on cryosurvival are not dependent on serum supplementation. Table 1.Effect of addition of l-carnitine during culture on post-thaw re-expansion and hatching rates

Isolation of soluble scFv antibody fragments specific for small biomarker molecule, L-Carnitine, using phage display

Isolation of single chain antibody fragment (scFv) clones from naïve Tomlinson I+J phage display libraries that specifically bind a small biomarker molecule, L-Carnitine, was performed using iterative affinity selection procedures. L-Carnitine has been described as a conditionally essential nutrient for humans. Abnormally high concentrations of L-Carnitine in urine are related to many health disorders including diabetes mellitus type 2 and lung cancer. ELISA-based affinity characterization results indicate that selectants preferentially bind to L-Carnitine in the presence of key bioselecting component materials and closely related L-Carnitine derivatives. In addition, the affinity results were confirmed using biophysical fluorescence quenching for tyrosine residues in the V segment. Small-scale production of the soluble fragment yielded 1.3mg/L using immunopure-immobilized protein A affinity column. Circular Dichroism data revealed that the antibody fragment (Ab) represents a folded protein that mainly consists of β-sheets. These novel antibody fragments may find utility as molecular affinity interface receptors in various electrochemical biosensor platforms to provide specific L-Carnitine binding capability with potential applications in metabolomic devices for companion diagnostics and personalized medicine applications. It may also be used in any other biomedical application where detection of the L-Carnitine level is important.

Probiotic supplementation and trimethylamine-N-oxide production following a high-fat diet.

The objective of this study was to test the hypothesis that the multi-strain probiotic VSL#3 would attenuate the increase in fasting plasma concentrations of trimethylamine-N-oxide (TMAO) following a high-fat diet. Nineteen healthy, non-obese males (18-30 years) participated in the present study. Following a 2-week eucaloric control diet, subjects were randomized to either VSL#3 (900 billion live bacteria) or placebo (cornstarch) during the consumption of a hypercaloric (+1,000 kcalday(-1) ), high-fat diet (55% fat) for 4 weeks. Plasma TMAO, L-carnitine, choline, and betaine (UPLC-MS/MS) were measured at baseline and following a high-fat diet. Plasma TMAO significantly increased 89% ± 66% vs. 115% ± 61% in both the VSL#3 and placebo groups, respectively; however, the magnitude of change in plasma TMAO was not different (P > 0.05) between them. Plasma L-carnitine, choline, and betaine concentrations did not increase following the high-fat diet in either group. A high-fat diet increases plasma TMAO in healthy, normal-weight, young males. However, VSL#3 treatment does not appear to influence plasma TMAO concentrations following a high-fat diet. Future studies are needed to determine whether other therapeutic strategies can attenuate the production of TMAO.

The Relationship Between Trimethylamine-N-Oxide and Prevalent Cardiovascular Disease in a Multiethnic Population Living in Canada

BackgroundMicroflora-dependent trimethylamine-N-oxide (TMAO) formation, which results from intake of choline and L-carnitine-rich food, shows promise as a predictor of cardiovascular disease (CVD) risk, but these associations have not been examined in ethnically diverse populations. In a multiethnic population-based study of adults in Canada, we assessed the stability of TMAO and L-carnitine in stored serum samples and their association with intimal medial thickness, prevalent risk factors, and clinical events. MethodsIn a randomly sampled cross-sectional study of 1286 Canadians, fasting serum samples were collected and stored. In 292 consecutive individuals (99 CVD cases and 193 unmatched control subjects), L-carnitine and TMAO concentrations were assessed using validated analytical approaches. ResultsThe mean (± SD) TMAO level was 1.998 ± 3.13 μM and L-carnitine was 42.29 ± 11.35 μM. The relative levels of the samples did not appreciably change after 3 freeze-thaw cycles (coefficient of variation, 5.6% and 4.7%, respectively). No significant association between L-carnitine levels and prevalent CVD was found, with adjustment for covariates (odds ratio, 1.57; 95% confidence interval, 0.58-4.26; P trend = 0.65), for highest vs lowest quintile group. TMAO levels showed a significant, graded association with prevalent CVD (odds ratio, 3.17; 95% confidence interval, 1.05-9.51; P trend = 0.02). After further adjustment for diabetes status, meat, fish, and cholesterol intake, the association remained significant. No significant association between carotid intimal medial thickness and L-carnitine (P = 0.64) or TMAO (P = 0.18) was found. ConclusionsSerum TMAO and L-carnitine analysis on stored samples is reliable. Our findings support an association between TMAO with prevalent CVD in a multiethnic population. This finding requires replication in larger studies in which dietary intake and stored serum samples exist.

Effect of L-carnitine supplementation on maturation and early embryo development of immature mouse oocytes selected by brilliant cresyle blue staining.

The present study was designed to investigate the effect of L-carnitine treatment during IVM on nuclear and cytoplasmic maturation of immature oocytes selected by Brilliant Cresyle Blue (BCB) staining, and their subsequent developmental competence. Compact cumulus-oocyte complexes (COCs) were collected from NMRI mice ovaries and stained with BCB staining. BCB+ (colored cytoplasm) oocytes were then cultured in tissue culture medium (TCM) 199 with 0.0, 0.3 and 0.6 mg/ml L-carnitine. The both L-carnitine concentrations significantly increased the intracellular glutathione (P<0.001), nuclear maturation (P<0.01) and expression levels of cyclin-dependent kinase1 (CDK1) (P<0.05). Moreover, treated oocytes with 0.6 mg/ml L-carnitine showed increased (P < 0.05) expression of mitogen-activated protein kinase1 (MAPK1) mRNA. Also, adding L-carnitine (0.6 mg/ml) to IVM medium significantly increased the cleavage rate (P<0.05). The blastocyst development rate (BDR) in the both L-carnitine treated groups was significantly higher (P<0.001) than the control group. L-carnitine had no significant effect on total blastocyst cell numbers. These data indicated that L-carnitine supplementation during IVM of immature BCB+ oocytes improved preimplantation developmental competence of oocytes after IVF, probably by accelerating cytoplasmic and nuclear maturation of oocytes. It may provide a novel approach to improving ART outcomes in infertile couples.

Protective effect of L-carnitine on Phenylalanine-induced DNA damage.

The pathogenesis and the progression of phenylketonuria (PKU), an inborn error of phenylalanine (Phe) metabolism, have been associated with oxidative damage. Moreover, it has been increasingly postulated the antioxidant properties of L-Carnitine (LC). The aim of this study was to verify the effect of LC on Phe-induced DNA damage. The in vitro effect of different concentrations of LC (15, 30, 120 and 150μM) on DNA damage-induced by high phenylalanine levels (1000 and 2500μM) was examined in white blood cells from normal individuals using the comet assay. Urinary 8-hydroxydeoguanosine (8-OHdG) levels, a biomarker of oxidative DNA damage, and plasmatic sulfhydryl content were measured in eight patients with classical PKU, under therapy with protein restriction and supplemented with a special formula containing LC, and in controls individuals. Both in vitro tested Phe concentrations (1000 and 2500μM) have resulted in DNA damage index significantly higher than control group. The in vitro co-treatment with Phe and LC reduced significantly DNA damage index when compared to Phe group. The urinary excretion of 8-OHdG and plasmatic sulfhydryl content presented similar levels in both groups analyzed (controls and treated PKU patients). In treated PKU patients, urinary 8-OHdG levels were positively correlated with blood Phe levels and negatively correlated with blood LC concentration and plasmatic sulfhydryl content. The present work yields experimental evidence that LC can reduce the in vitro DNA injury induced by high concentrations of phenylalanine, as well as, allow to hypothesize that LC protect against DNA damage in patients with PKU.

L-carnitine affects osteoblast differentiation in NIH3T3 fibroblasts by the IGF-1/PI3K/Akt signalling pathway.

Fibroblasts in soft tissues are one of the progenitors of ectopic calcification. Our previous experiment found that the serum concentrations of small metabolite L-carnitine (LC) decreased in an ectopic calcification animal model, indicating LC is a potential calcification or mineralization inhibitor. In this study, we investigated the effect of LC on NIH3T3 fibroblast osteoblast differentiation, and explored its possible molecular mechanisms. Two concentrations of LC (10 μM and 100 μM) were added in Pi-induced NIH3T3 fibroblasts, cell proliferation was compared by MTT assays, osteoblast differentiation was evaluated by ALP activity, mineralized nodules formation, calcium deposition, and expressions of the osteogenic marker genes. Our results indicated that 10 μM LC increased the proliferation of NIH3T3 cells, but 100 μM LC slightly inhibited cell proliferation. 100 μM LC inhibits NIH3T3 differentiation as evidenced by decreases in ALP activity, mineralized nodule formation, calcium deposition, and down-regulation of the osteogenic marker genes ALP, Runx2 and OCN, meanwhile 10 μM of LC exerts an opposite effect that promotes NIH3T3 osteogenesis. Mechanistically, 100 μM LC significantly inhibits IGF-1/PI3K/Akt signalling, while 10 μM LC slightly activates this pathway. Our study suggests that a decease in LC level might contribute to the development of ectopic calcification in fibroblasts by affecting IGF-1/PI3K/Akt, and addition of LC may benefit patients with ectopic calcification.

Determination of L-carnitine in milk and dairy products by hydrophilic liquid chromatography-tandem mass spectrometry

An analytical method was developed for the determination of L-carnitine in milk and dairy products using hydrophilic interaction chromatography-tandem mass spectrometry (HILIC-MS/MS). The samples were extracted with 2% (v/v) acetic acid solution, and the protein was precipitated with acetonitrile subsequently. The separation of L-carnitine was carried out on an Acquity UPLC BEH HILIC column using ammonium acetate and acetonitrile as mobile phases. The quantification analysis of the target compound was performed under multiple reaction monitoring (MRM) mode by external standard method. A good linear relationship was obtained between the peak area and concentration of L-carnitine in the range of 1-100 µg/L with the correlation coefficient more than 0.99. The limit of quantification (LOQ ) of L-carnitine was 0. 01 mg/kg. The spiked recoveries were 96.0%-103.4%. The precisions (RSDs ) were 1.2%-4.3%. The sample preparation was simple and rapid, and the results were precise and sensitive. The developed method is suitable for the study of concentration of L-carnitine in milk and dairy products, and the technical support for the infant formula is provided.

Validation of a Stability-Indicating RP-HPLC Method for Determination of l-Carnitine in Tablets.

A rapid and stability-indicating RP-HPLC method was developed for determination of l-carnitine in tablets. The separation was based on a C18 analytical column using a mobile phase which consisted of 0.05 M phosphate buffer (pH = 3): ethanol (99 : 1), including 0.56 mg/mL of sodium 1-heptanesulfonate. Column temperature was set at 50°C and quantitation was achieved by UV detection at 225 nm. In forced degradation studies, the drug was subjected to oxidation, hydrolysis, photolysis, and heat. Among the different stress conditions, the exposure to acidic and basic conditions was found to be an important adverse stability factor. The method was validated for specificity, selectivity, linearity, precision, accuracy, and robustness. The applied procedure was found to be linear in l-carnitine concentration range of 84.74–3389.50 µg/mL (r 2 = 0.9997). Precision was evaluated by replicate analysis in which relative standard deviation (RSD) values for areas were found below 2.0%. The recoveries obtained (100.83%–101.54%) ensured the accuracy of the developed method. The expanded uncertainty (3.14%) of the method was also estimated from method validation data. Accordingly, the proposed validated and rapid procedure was proved to be suitable for routine analyzing and stability studies of l-carnitine in tablets.

L-Carnitine: a new insight into the pathogenesis of endometrial cancer.

The present study aims to specify the role of L-carnitine in the pathogenesis of endometrial cancer by comparing the serum total L-carnitine levels of endometrial cancer patients with those of healthy women. Serum total L-carnitine concentrations were measured in patients with endometrioid-type endometrial cancer (n = 20) and healthy controls (n = 20) who were matched with respect to age and body mass index (BMI). Stage I endometrial cancer was diagnosed in 12 women (60.0%) whereas three women (15.0%) had stage II disease, three women (15.0%) had stage III disease and two women (10.0%) had stage IV disease. The healthy controls and endometrial cancer patients were statistically similar in aspect of age, gravidity, parity, BMI, waist-to-thigh ratio, waist-to-hip ratio, menopause, complete blood count parameters, and serum biochemistry. Serum total L-carnitine levels of women with endometrial cancer were significantly lower than those of healthy women (respectively, 5,519.4 ± 2,712.5 vs 7,940.8 ± 3,566.6 ng/dl, p = 0.021). Moreover, serum total L-carnitine levels decreased significantly and progressively with advancing stage (stage I vs II vs III vs IV; 6,294.0 ± 2,885.1 vs 5,800.0 ± 441.2 vs 4,016.0 ± 2,833.3 vs 2,560.0 ± 67.9 ng/dl; p = 0.021). This is the first study to hypothesize that L-carnitine deficiency participates in the pathogenesis of endometrial cancer by means of a mechanism which is unrelated with obesity and increased amount of fat in human body.

The influence of chronic L-carnitine supplementation on the formation of preneoplastic and atherosclerotic lesions in the colon and aorta of male F344 rats.

l-Carnitine, a key component of fatty acid oxidation, is nowadays being extensively used as a nutritional supplement with allegedly “fat burning” and performance-enhancing properties, although to date there are no conclusive data supporting these claims. Furthermore, there is an inverse relationship between exogenous supplementation and bioavailability, i.e., fairly high oral doses are not fully absorbed and thus a significant amount of carnitine remains in the gut. Human and rat enterobacteria can degrade unabsorbed l-carnitine to trimethylamine or trimethylamine-N-oxide, which, under certain conditions, may be transformed to the known carcinogen N-nitrosodimethylamine. Recent findings indicate that trimethylamine-N-oxide might also be involved in the development of atherosclerotic lesions. We therefore investigated whether a 1-year administration of different l-carnitine concentrations (0, 1, 2 and 5 g/l) via drinking water leads to an increased incidence of preneoplastic lesions (so-called aberrant crypt foci) in the colon of Fischer 344 rats as well as to the appearance of atherosclerotic lesions in the aorta of these animals. No significant difference between the test groups regarding the formation of lesions in the colon and aorta of the rats was observed, suggesting that, under the given experimental conditions, l-carnitine up to a concentration of 5 g/l in the drinking water does not have adverse effects on the gastrointestinal and vascular system of Fischer 344 rats.Electronic supplementary materialThe online version of this article (doi:10.1007/s00204-014-1341-4) contains supplementary material, which is available to authorized users.

Effective dosing of L-carnitine in the secondary prevention of cardiovascular disease: a systematic review and meta-analysis.

BackgroundL-carnitine supplementation has been associated with a significant reduction in all-cause mortality, ventricular arrhythmia, and angina in the setting of acute myocardial infarction (MI). However, on account of strict homeostatic regulation of plasma L-carnitine concentrations, higher doses of L-carnitine supplementation may not provide additional therapeutic benefits. This study aims to evaluate the effects of various oral maintenance dosages of L-carnitine on all-cause mortality and cardiovascular morbidities in the setting of acute MI.MethodsAfter a systematic review of several major electronic databases (PubMed, EMBASE, and the Cochrane Library) up to November 2013, a meta-analysis of five controlled trials (n = 3108) was conducted to determine the effects of L-carnitine on all-cause mortality and cardiovascular morbidities in the setting of acute MI.ResultsThe interaction test yielded no significant differences between the effects of the four daily oral maintenance dosages of L-carnitine (i.e., 2 g, 3 g, 4 g, and 6 g) on all-cause mortality (risk ratio [RR] = 0.77, 95% CI [0.57-1.03], P = 0.08) with a statistically insignificant trend favoring the 3 g dose (RR = 0.48) over the lower 2 g dose (RR = 0.62), which was favored over the higher 4 g and 6 g doses (RR = 0.78, 0.78). There was no significant differences between the effects of the daily oral maintenance dosages of 2 g and 6 g on heart failure (RR = 0.53, 95% CI [0.25-1.13], P = 0.10), unstable angina (RR = 0.90, 95% CI [0.51-1.58], P = 0.71), or myocardial reinfarction (RR = 0.74, 95% CI [0.30-1.80], P = 0.50).ConclusionsThere appears to be no significant marginal benefit in terms of all-cause mortality, heart failure, unstable angina, or myocardial reinfarction in the setting of acute MI for oral L-carnitine maintenance doses of greater or less than 3 g per day.

Trimethylamine generation in patients receiving hemodialysis treated with l-carnitine.

Hepatic biotransformation of gut microbiota-derived trimethylamine (TMA) to trimethylamine N-oxide (TMAO) may enhance cardiovascular risk [1, 2]. The source of TMA is fish-derived TMAO [3], meat-derived betaine/choline/phosphatidylcholine [1, 2], and the medicine l-carnitine [4]. There is no clear information on the metabolic fate of TMA/TMAO in patients receiving hemodialysis and treated with l-carnitine orally and intravenously. We report data showing that plasma TMA/TMAO concentrations in hemodialysis patients may be in the same range as those of healthy subjects consuming a typical diet in Japan. Fourteen subjects, randomly selected from a cohort of 18 male and 6 female Japanese patients (47–89 years) with diabetes mellitus or chronic glomerulonephritis who had undergone hemodialysis for several years, were divided into five untreated controls, five treated with 1000 mg of l-carnitine (l-Cartin injection, Otsuka, Tokyo, Japan) intravenously on 3 days (Days 1, 3 and 5) over 1 week, and four treated with 300 mg of oral l-carnitine (l-Cartin tablets, Otsuka) three times per day for 7 days. Ten healthy volunteers (22–59 years) were also enrolled. This study was approved by the ethics committees of Minami Ikebukuro Clinic and Showa Pharmaceutical University. Total carnitine and TMA/TMAO concentrations were determined by Kainos laboratories (Tokyo, Japan) and by gas chromatography [3]. Carnitine concentrations for 2–8 weeks after treatment with l-carnitine were significantly higher than those in the untreated group (Figure 1). Plasma TMA concentrations after oral treatment with l-carnitine were higher than those of the untreated group. TMAO concentrations did not increase significantly after treatments and were in the range of daily interindividual variations. In separate observations in 22 of the 24-patient cohort, pre-dialysis plasma concentrations of carnitine (33 ± 4 µM, mean ± SD), TMA (34 ± 20 µM) and TMAO (189 ± 94 µM) were decreased by hemodialysis (12 ± 8, 25 ± 8 and 140 ± 77 µM, respectively). These concentration ranges were comparable with TMA (26 ± 17 µM) and TMAO (197 ± 100 µM) in healthy volunteers (n = 10). The present plasma levels of TMA and/or TMAO in Japanese patients were one or two orders of magnitude higher than the reported concentrations in Caucasians [1, 2], but were consistent with a review of patients undergoing hemodialysis [4]. Fig. 1. Plasma concentrations of total l-carnitine (A), TMA (B) and TMAO (C) in untreated patients (circles) and in patients treated with l-carnitine intravenously (triangles) and orally (squares). The three bars are averages of the three groups (n = 4 or 5). ... Some TMA formation in a Japanese cohort treated with l-carnitine orally was confirmed, but the levels of TMA/TMAO both pre-/post-dialysis in patients not receiving l-carnitine were in the same ranges as those of the control subjects.

Urine l-carnitine excretion in hypertensive adolescents

AimThis study was performed to test the hypothesis that urinary levels of l-carnitine and its derivatives are enhanced in children and adolescents with hypertension and also check if analyzed parameters may serve as early markers of subclinical renal damage.MethodsThe study included 112 children and adolescents (67 males and 45 females) aged median 10–18 years. Participants were divided into two groups: HT—64 subjects with confirmed primary hypertension and R—reference group—48 subjects with white-coat hypertension.Urinary Free and Total l-carnitine were determined by the enzymatic method of Cederblad. The l-carnitine levels were expressed as urinary ratio in micromole per gram creatinine (μmol/g).ResultsThe urinary excretion of Total and Free l-carnitine was significantly higher in hypertensive adolescents in comparison to reference group—white coat hypertension. Other important findings were positive correlations between Free l-carnitine/cr., Total l-carnitine/cr. ratio and serum uric acid level, serum cholesterol level and systolic blood pressure.ConclusionThe results of this study do not explain the increased urine levels of l-carnitine. The most likely reason for excessive urinary loss was disturbed renal tubular reabsorption. It is possible to hypothesize that in hypertensive adolescents subclinical kidney dysfunction occurs. It is proposed that studies examining the concurrent plasma and urine concentration of l-carnitine and correlation with acknowledged proximal tubular markers are needed.

Mildronate, the inhibitor of l-carnitine transport, induces brain mitochondrial uncoupling and protects against anoxia-reoxygenation

The preservation of mitochondrial function is essential for normal brain function after ischaemia-reperfusion injury. l-carnitine is a cofactor involved in the regulation of cellular energy metabolism. Recently, it has been shown that mildronate, an inhibitor of l-carnitine transport, improves neurological outcome after ischaemic damage of brain tissues. The aim of the present study was to elucidate the mitochondria targeted neuroprotective action of mildronate in the model of anoxia-reoxygenation-induced injury. Wistar rats were treated daily with mildronate (per os; 100mg/kg) for 14 days. The acyl-carnitine profile was determined in the brain tissues. Mitochondrial respiration and the activities of carnitine acetyltransferase (CrAT) and tricarboxylic acid (TCA) cycle enzymes were measured. To assess tolerance to ischaemia, isolated mitochondria were subjected to anoxia followed by reoxygenation. The mildronate treatment significantly reduced the concentrations of free l-carnitine (FC) and short-chain acyl-carnitine (AC) in brain tissue by 40–76%, without affecting the AC:FC ratio. The activities of CrAT and TCA cycle enzymes were slightly increased after mildronate treatment. Despite partially induced uncoupling, mildronate treatment did not affect mitochondrial bioenergetics function under normoxic conditions. After exposure to anoxia-reoxygenation, state 3 respiration and the respiration control ratio were higher in the mildronate-treated group. The results obtained demonstrate that mildronate treatment improves tolerance against anoxia-reoxygenation due to an uncoupling preconditioning-like effect. Regulating l-carnitine availability provides a potential novel target for the treatment of cerebral ischaemia and related complications.