Odd-numbered Long-chain Fatty Acids Research Articles

Plasmonic Hydroxyl Radical-Driven Epoxidation of Fatty Acid Double Bonds in Nanoseconds for On-Tissue Mass-Spectrometric Analysis and Bioimaging.

Lipids represent a large family of compounds with highly diverse structures that are involved in complex biological processes. A photocatalytic technique of on-tissue epoxidation of C=C double bonds has been developed for in situ mass spectrometric identification and spatial imaging of positional isomers of lipids. It is based on the plasmonic hot-electron transfer from irradiated gold nanowires to redox-active organic matrix compounds that undergo bond cleavages and generate hydroxyl radicals in nanoseconds. Intermediate radical anions and negative fragment ions have been unambiguously identified. Under the irradiation of a pulsed laser of the third harmonic of Nd3+:YAG (355 nm), the hydroxyl radical-driven epoxidation of unsaturated lipids with different numbers of C=C bonds can be completed in nanoseconds with high yields of up to 95%. Locations of C=C bonds were recognized with diagnostic fragment ions that were produced by either collision with an inert gas or auto-fragmentation resulting from the impact of energetic hot electrons and vibrational excitation. This technique has been applied to the analysis of breast cancer tissues of mice models without extensive sample processes. It was experimentally demonstrated that C=C bonds may be formed at different positions of not only regular mono- or poly-unsaturated fatty acids but also other odd-numbered long-chain fatty acids.

Long-term follow-up with filter paper samples in patients with propionic acidemia.

BackgroundPropionic acidemia (PA) is an inherited disorder caused by deficiency of propionyl CoA carboxylase. Most patients with this disorder are diagnosed during the neonatal period because of severe metabolic acidosis and hyperammonemia. Patients are required to undergo blood and urine analysis at least 3 to 4 times per year, depending on age and metabolic control.MethodsWe designed a prospective study in which we investigated the results from blood and urinary samples collected monthly in filter paper from 10 PA patients followed in a single metabolic reference center from January 2015 to September 2017. The aim of this study was to evaluate the usefulness of filter paper samples in the follow‐up of the PA patients.ResultsDuring the follow‐up period, 163 dried blood spot (DBS) and 119 urine dried spot samples were analyzed and compared with 160 plasma and 103 liquid urine specimens; 64 specimens of plasma were analyzed for odd‐numbered long‐chain fatty acids (OLCFAs). A total of 40 metabolic crises, 18 of them with hyperammonemia were documented. We observed a strong correlation between the filter paper and the urine/plasma samples for the main PA parameters both in stable metabolic conditions as well as in acute decompensations. Also, there was a strong correlation between OLCFAs measured in plasma and quantification of odd number acylcarnitines in DBS.ConclusionsWe conclude that filter paper blood and urinary samples can be used for the follow‐up of the patients with PA, correctly reflecting their metabolic situation.

Methylmalonic acidaemia: Examination of genotype and biochemical data in 32 patients belonging to mut, cblA or cblB complementation group

Methylmalonic acidaemia (MMA) is a genetic disorder caused by defects in methylmalonyl-CoA mutase or in any of the different proteins involved in the synthesis of adenosylcobalamin. The aim of this work was to examine the biochemical and clinical phenotype of 32 MMA patients according to their genotype, and to study the mutant mRNA stability by real-time PCR analysis. Using cellular and biochemical methods, we classified our patient cohort as having the MMA forms mut (n = 19), cblA (n = 9) and cblB (n = 4). All the mut (0) and some of the cblB patients had the most severe clinical and biochemical manifestations, displaying non-inducible propionate incorporation in the presence of hydroxocobalamin (OHCbl) in vitro and high plasma odd-numbered long-chain fatty acid (OLCFA) concentrations under dietary therapy. In contrast, mut (-) and cblA patients exhibited a milder phenotype with propionate incorporation enhanced by OHCbl and normal OLCFA levels under dietary therapy. No missense mutations identified in the MUT gene, including mut (0) and mut (-) changes, affected mRNA stability. A new sequence variation (c.562G>C) in the MMAA gene was identified. Most of the cblA patients carried premature termination codons (PTC) in both alleles. Interestingly, the transcripts containing the PTC mutations were insensitive to nonsense-mediated decay (NMD).

Odd-numbered long-chain fatty acids in erythrocyte phospholipids as long-term follow-up parameter in propionic acidemia

Odd-numbered long-chain fatty acids (OLCFAs) increase in total erythrocyte lipids or plasma in patients with propionic acidemia and have been proposed as a useful parameter of metabolic control. However, up to now no parameter for long-term metabolic control has been available for this disorder. In contrast to previous investigations, we investigated OLCFAs with particular reference to differences for two phospholipid fractions of erythrocyte lipids. The phosphatidylethanolamine (PE) fraction of erythrocyte lipids has a slow turnover rate and might reflect the metabolic propionyl-CoA burden over a period of some weeks. Distinct increases in OLCFA of up to 200% in the phosphatidylcholine fraction were observed after episodes of metabolic decompensation and maximum OLCFA levels were found 3 weeks after the onset of a crisis. As a sign of poor metabolic control, OLCFA levels in PE steadily increased after recurrent episodes of decompensation, but not in clinically stable patients. However, in one patient with clinically good metabolic control, OLCFAs continually increased after the second year of life for an as yet unknown reason. In conclusion, data from this study indicate the usefulness of OLCFA analysis in the PE fraction of erythrocyte lipids, in particular with regard to the need for such a parameter for evidence-based improvement of management strategies in propionic acidemia.

Odd-numbered long-chain fatty acids in propionic acidaemia.

In patients with propionic acidaemia (PA), the increased intracellular concentration of propionyl-CoA leads to a relative abundance of odd-numbered long-chain fatty acids (OLCFAs) in body lipids. We investigated the relative amount of OLCFA in erythrocyte membrane lipids over a period of 1-8 years in five patients with early onset PA and present their clinical outcome. After extraction from erythrocyte membrane lipids and esterification, fatty acids were analysed by capillary column gas chromatography. The sum of the OLCFA 15- and 17- carbon saturated and 17-carbon monounsaturated fatty acids (C15:0, C17:0, C17:1) was calculated and expressed as a percentage of the total C14-C22 fatty acids in the sample. Three patients (pccBC-complementation group) presented with a stable clinical course and showed OLCFA values usually below 1.9% (median % +/- SD: 1.4+/-0.5, 1.6+/-0.5, 1.8+/-0.5). Two patients (pccA-complementation group) had a more severe course of the disease and showed higher medians and a broader range of OLCFA levels (2.2+/-1.2 and 2.2+/-0.8). Our study shows that odd-numbered long-chain fatty acid concentrations are increased in patients with propionic acidaemia and are higher in those with a more severe clinical course. The value of odd-numbered long-chain fatty acids in the assessment of the phenotypic severity and in the management of propionic acidaemia remains to be proven in a prospective long-term study with more patients of differing phenotype.

Odd-Numbered Long-Chain Fatty Acids (OLCFA) in Erythrocyte Membrane Lipids as Follow-up Parameter in Propionic Acidemia 165

Odd-Numbered Long-Chain Fatty Acids (OLCFA) in Erythrocyte Membrane Lipids as Follow-up Parameter in Propionic Acidemia 165

Plasma total odd-chain fatty acids in the monitoring of disorders of propionate, methylmalonate and biotin metabolism.

Total plasma odd-numbered long-chain fatty acids were analysed in patients with methylmalonic acidaemia (vitamin B12-responsive and unresponsive), combined methylmalonic acidaemia/homocystinuria (CblC), propionic acidaemia (both neonatal-onset and late-onset), biotinidase deficiency and holocarboxylase synthase deficiency, as well as in hospital controls. Total odd-numbered long-chain fatty acids (C15:0, C17:1 and C17:0) were expressed as a percentage of total C12-C20 fatty acids. Control values were 0.72% +/- 0.31% (n = 12). Normalization of the percentage of odd-chain fatty acids occurred in all vitamin-responsive patients, following the institution of vitamin treatment. In general the neonatal-onset propionic acidaemia and B12-unresponsive methylmalonic acidaemia patients had the highest plasma odd-chain fatty acid concentrations, which correlated with the clinical condition but not with the urinary excretion of methylcitrate or methylmalonate. Plasma odd-chain fatty acid concentrations and methylmalonate excretions in CblC patients reacted very well to vitamin B12 treatment, but with no clinical response. Measurement of plasma odd-chain fatty acids is of no value for the monitoring of defects of biotin metabolism.

Contribution of odd-numbered fatty acid oxidation to propionate production in neonates with methylmalonic an propionic acidaemias

Newborns with disturbed propionate metabolism (propionic and methylmalonic acidaemias) accumulate during fetal life large amounts of odd-numbered long-chain fatty acids in adipose tissue (8% to 10% of total fatty acids vs 1% in normal controls) and other body lipids. During periods of acute catabolism, such as in the early days of life, mobilization and oxidation of odd-chain fatty acids from adipose tissue yield extensive amounts of toxic propionyl-CoA in the mitochondria and thus might contribute to the severe illness of the patients. Our data suggest that not only catabolism of protein but also that of adipose tissue has to be avoided in order to alleviate the neonatal illness of patients with propionic and methylmalonic acidaemias. It seems prudent to avoid prolonged fasting during the 1st year of life.

ODD-NUMBERED LONG-CHAIN FATTY ACIDS IN ERYTHROCYTE MEMBRANE LIPIDS AND PROPIONIC ACID IN PLASMA FOR METABOLIC CONTROL OF PATIENTS WITH PROPIONIC ACIDAEMIA

In patients with propionic acidaemia (PA) and methylmalonic acidaemia (MMA) the increased concentration of propionyl-CoA in cells leads to a relative abundance of odd-numbered long-chain fatty acids (OLCFA) in body lipids. During a period of 5 years we used the OLCFA content in erythrocyte membranes for long-term metabolic control in 5 patients with PA. The determination of OLCFA in erythrocyte membrane lipids was done by capillary column gas liquid chromatography and the sums of the individual OLCFA- C15:0, C17:0 and C17:1- were calculated and expressed as a percentage of the total C14-C22 fatty acids. In 3 patients OLCFA are usually below 1.9% indicating a good metabolic control. In one patient, metabolic control is fair (OLCFA < 3.1%) and one neonate with severe course of PA showed a decrease of OLCFA with age and an increase during metabolic decompensation before death. In general, during acute metabolic crisis we only found an elevation of propionic acid in plasma. After crisis there was a delayed increase of OLCFA. In one patient a decrease of OLCFA could be seen after therapy with metronidazole. OLCFA seem to be a valuable parameter for long-term metabolic control in patients with PA.

Accumulation of Odd-Numbered Long-Chain Fatty Acids in Fetuses and Neonates with Inherited Disorders of Propionate Metabolism

Fetuses affected with propionic acidemia incorporate great amounts of odd-numbered long-chain fatty acids (OLCFA) into their body lipids. This is due to abundant supply with precursor amino acids of propionyl-CoA throughout pregnancy. After birth, the lower provision of precursor amino acids during dietary treatment compared with fetal life results in a decline of propionyl-CoA production and therefore OLCFA synthesis. However, the observed decrease of OLCFA may also partly reflect the recovery from acute ketoacidotic episodes that the patients experienced soon after birth as long as they were undiagnosed. In a patient with vitamin B12-responsive methylmalonic aciduria treated prenatally with large doses of vitamin B12 given to the mother, the cord plasma lipids contained normal amounts of OLCFA. This indicates that prenatal therapy led to an increased flux of propionyl-CoA through the defective methylmalonyl-CoA mutase step. Thus, in addition to the quantification of a decline in methylmalonic acid in maternal urine, OLCFA in cord blood lipids might be a further parameter for evaluating prenatal treatment in patients with vitamin B12-responsive methylmalonic aciduria.

Abnormality of odd-numbered long-chain fatty acids in erythrocyte membrane lipids from patients with disorders of propionate metabolism.

In propionic acidemia and methylmalonic aciduria, disorders of propionate catabolism, excess intracellular propionyl-CoA leads to an enhanced synthesis of odd-numbered long-chain fatty acids. Red cell membrane lipids of patients with these disorders contain pentadecanoic, heptadecanoic, and heptadecenoic acids in excess of normal levels. The odd-numbered long-chain fatty acid content may reflect individual differences in severity of these disorders as well as the degree of effective dietary control. It might be a useful long term parameter for determining the effectiveness of clinical management.

Odd-numbered long-chain fatty acid contents in erythrocyte membrane phospholipids in patients with an impaired propionate utilization.

Patients with the inherited disorders of propionate and methylmalonate metabolism such as propionic acidaemia (PA; McKusick 23200) and methylmalonic acidurias (MMA; McKusick 25100) accumulate odd-numbered long-chain fatty acids (OLCFA) in lipids of various organs (Hommes et al., 1968; Gompertz, 1970; Gompertz et al., 1970; Kishimoto et al., 1973). The accumulation of OLCFA also occurs in cultured skin fibroblasts of PA and MMA patients (Giudici et al., 1986). This is due to the fact that intracellularly accumulated propionyl-CoA can replace acetyl-CoA as a ‘primer’ for long-chain fatty acid synthesis.

The occurrence of β-hydroxy- n-valeric acid in a patient with propionic and methylmalonic acidemia

1. 1. The patient, a boy, was suffering from episodes of vomiting, dystrophy, muscular hypotonia and weakness, psychomotoric retardation, respiratory tract infections, and attacks of metabolic acidosis from the first days of life until death at the age of 10 months. 2. 2. A small amount of serum (1.5 ml), obtained by heart puncture shortly after death, was the only material left from the patient. The serum contained 3.5 mmol/l of propionic acid, 1.13 mM of methylmalonic acid, and large amounts of β-hydroxybutyrate and lactate. In addition, the analyses revealed 1.54 mM of β-hydroxy- n-valeric acid, a compound not detected in human material before. The fatty acid pattern showed increased amounts of the odd-numbered pentadecanoic acid and heptadecanoic acid. There were elevated amounts of lysine, leucine, isoleucine and valine, while the other amino acids were within the normal range. 3. 3. The clinical picture as well as the biochemical findings leave no doubt that the patient suffered from a metabolic disorder, most likely an inborn error of metabolism. The primary defect of the patient was most likely located to the conversion of methylmalonyl CoA to succinyl CoA. Since accumulation of propionate has not been described in patients with a defective methylmalonyl-CoA mutase, the patient may possibly have a defect located to another enzyme in this metabolic sequence, probably the methylmalonyl CoA racemase. The β-hydroxy- n-valeric acid and the odd-numbered long-chain fatty acids are likely to stem from the accumulated propionyl CoA, which may compete with acetyl-CoA in several condensing reactions.