Unraveling the pathway of lipoic acid biosynthesis

Lipoic acid is almost universally required for aerobic metabolism. However, the mechanism for its synthesis and incorporation into proteins has remained elusive. A groundbreaking study published in the December issue of Chemistry & Biology uncovers critical features of the lipoic acid biosynthetic pathway.     

Thermodynamic analysis of lipoic acid crystallized with additives

Differential scanning calorimetry (DSC) was used as a screening technique to study the interaction between lipoic acid and amino acids. The samples containing lipoic acid and amino acids were prepared by two different methods (milling method and cooling crystallization method). Firstly, cooling method was used to prepare the sample with amino acids as additives. DSC results indicated that only phenylalanine has some interaction with lipoic acid. Further, in order to study the interaction between lipoic acid and phenylalanine, another method (milling method) was used to prepare the sample containing phenylalanine, and the effect of sample preparation methods was studied through IR analysis and the X-ray powder diffraction. On the basis of above researches, the possible interaction mechanisms between lipoic acid and phenylalanine were proposed.     

硫辛酸“结构-抗氧化性”关系的理论研究

采用密度泛函方法,以键解离焓(EBD)和电离势(EIP)为理论指标评价了硫辛酸清除自由基的活性,发现硫辛酸的EBD和EIP均小于具有抗氧化活性的VC、VE和没食子酸.结合优势构象的几何构型、电荷分布和前线轨道的布居分析结果,发现二硫五元环是分子中的活性区域,电荷密度较大,可以与金属离子螯合;S—S键较长,且EBD和EIP均较小,可以形成自由基而发挥抗氧化作用.羧基部分可以调节分子的水溶性,且电荷密度较大,也可以和金属离子螯合.初步明确了硫辛酸的"结构-抗氧化性"关系.     

Nanoparticles of lipoic acid esters: preparation and antioxidant effect

Five lipoic acid esters were synthesized, and their nanoforms based on copolymer Pluronic F68 and phosphatidylcholine were obtained. The most pronounced antioxidant effect on neutrophils activated by phorbol-12-myristate-13-acetate was shown by the nanodispersion containing the derivative of lipoic acid with hexane-1,6-diol, reducing the concentration of reactive oxygen species by 7.5 times.     

Mechanistic investigations of lipoic acid biosynthesis in Escherichia coli: both sulfur atoms in lipoic acid are contributed by the same lipoyl synthase polypeptide

Lipoyl synthase catalyzes the final step in the de novo biosynthesis of the lipoyl cofactor, which is the insertion of two sulfur atoms into an octanoyl chain that is bound in an amide linkage to a conserved lysine on a lipoyl-accepting protein. We show herein that the sulfur atoms in the lipoyl cofactor are derived from lipoyl synthase itself, and that each lipoyl synthase polypeptide contributes both of the sulfur atoms to the intact cofactor.     

Meldrum's acid and its analogs

2-Substituted or 2,2-disubstituted 4,6-dioxo-1,3-dioxanes, which are analogs of Meldrum's acid and were obtained by reaction of malonic acid with acetic anhydride and the appropriate carbonyl compound in the presence of sulfuric acid, exist in solution preferably in a single form according to the PMR and ¹³C NMR spectral data. Fluctuation of the CH₂ group is not observed. The mass spectra are characteristic and are distinguished by splitting out of the carbonyl compound from the molecular ion or successive splitting out of CO₂ and ketene molecules. The Meldrum acid analogs do not affect plant growth but have a depressive effect on the central nervous system of animals and low toxicity.Translated from Khimiya Geterotsiklicheskikh Soedinenii, No. 11, pp. 1482–1486, November, 1975.     

Polynuclear tetrazole-containing amino acid analogs

New analogs of (D,L)-phenylalanine containing tetrazole rings were synthesized. The acidity constants of (D, L)-phenylalanine and (D,L)-tryptophane derivatives containing a tetrazole ring with no substituent on N¹ (pK _a = 3.0–3.1) and of the corresponding carboxylic acids (pK _a = 2.9–3.3) in aqueous methanol were determined by potentiometric titration.Translated from Zhurnal Organicheskoi Khimii, Vol. 40, No. 10, 2004, pp. 1576–1579.Original Russian Text Copyright © 2004 by Morozova, Esikov, Zubarev, Malin, Ostrovskii.     

Determination of lipoic acid in human urine by capillary zone electrophoresis

Fast, simple, and accurate CE method enabling determination of lipoic acid (LA) in human urine has been developed and validated. LA is a disulfide‐containing natural compound absorbed from the organism's diet. Due to powerful antioxidant activity, LA has been used for prevention and treatment of various diseases and disorders, e.g. cardiovascular diseases, neurodegenerative disorders, and cancer. The proposed analytical procedure consists of liquid–liquid sample extraction, reduction of LA with tris(2‐carboxyethyl)phosphine, derivatization with 1‐benzyl‐2‐chloropyridinium bromide (BCPB) followed by field amplified sample injection stacking, capillary zone electrophoresis separation, and ultraviolet‐absorbance detection of LA‐BCPB derivative at 322 nm. Effective baseline electrophoretic separation was achieved within 6 min under the separation voltage of 20 kV (∼80 μA) using a standard fused‐silica capillary (effective length 51.5 cm, 75 μm id) and BGE consisted of 0.05 mol/L borate buffer adjusted to pH 9. The experimentally determined limit of detection for LA in urine was 1.2 μmol/L. The calibration curve obtained for LA in urine showed linearity in the range 2.5–80 μmol/L, with R ² 0.9998. The relative standard deviation of the points of the calibration curve was lower than 10%. The analytical procedure was successfully applied to analysis of real urine samples from seven healthy volunteers who received single 100 mg dose of LA.     

Electron transfer from triplet thymine and thymidine to lipoic acid

Electron transfer from the triplet excited state of thymine or thymidine to the disulphide compound lipoic acid (RSSR) was studied using KrF laser flash photolysis (248 nm, 20 ns). The electron transfer reaction rate constants, measured at 310 nm, were determined to be 1.3×10¹⁰ M⁻¹ s⁻¹ and 6.9×10⁹ M⁻¹ s⁻¹ for thymine and thymidine respectively. The transient absorbance at 400 nm in the presence of the quencher is attributed to the anion radical of lipoic acid.     

The issue of HPLC determination of endogenous lipoic acid in human plasma

Lipoic acid (LA) is used extensively as a therapeutic agent for the treatment of various diseases. Many methods have been reported for the determination of LA plasma levels and its metabolites after its supplementation, but available information concerning endogenous plasma levels is still scarce. Studies which directly focused on determining the endogenous plasma levels provided highly controversial results, <4.9 nmol/L or 143.7–197.0 nmol/L. The main aim of this study was to verify the levels of free LA in the plasma of 40 individuals (17 women, 23 men). This group was nonsupplemented with LA and met the conditions for incorporation into the blood donors register. We measured the levels of LA using an HPLC method with very sensitive coulometric detection after previous sample preparation including deproteination and solid‐phase extraction with a Phenyl cartridge. Our limit of detection was 1.85 nmol/L and was better than the values reported in studies that directly focused on determining the endogenous plasma levels of LA: 2.4 and 4.9 nmol/L respectively. However, the levels of free LA in the plasma of nonsupplemented voluntary blood donors were not detectable in all cases. The presented results of our study show that endogenous concentrations of LA are <1.85 nmol/L.     

Synthesis of analogs of lysergic acid diethylamide

Conclusions A number of N-methyl-N[(-(-Indolyl)ethyl]glycine amides, representing analogs of lysergic acid diethylamide, were obtained.Translated from Izvestiya Akademii Nauk SSSR, Seriya Khimicheskaya, No. 11, pp. 2613–2614, November, 1972.     

Thermal analysis of lipoic acid and evaluation of the compatibility with excipientes

Differential scanning calorimetry (DSC) was used as a screening technique for assessing the compatibility of lipoic acid with some currently employed cosmetic excipients. In the first phase of the study DSC was used as a tool to detect any interaction. Based on the DSC results alone, methyl p-hydroxybenzoate, propyl p-hydroxybenzoate, butylated hydroxytoluene, non ionic self emulsifying wax, propylene glycol and acetylated lanolin were found to exhibit interaction with lipoic acid. Stressed binary mixtures (stored at 50 °C for 1 week) of lipoic acid and excipients were evaluated by HPLC. Binary mixtures were evaluated by IR spectroscopy.     

Assembly of the covalent linkage between lipoic acid and its cognate enzymes

Lipoic acid is synthesized from octanoic acid by insertion of sulfur atoms at carbons 6 and 8 and is covalently attached to a pyruvate dehydrogenase (PDH) subunit. We show that sulfur atoms can be inserted into octanoyl moieties attached to a PDH subunit or a derived domain. Escherichia coli lipB mutants grew well when supplemented with octanoate in place of lipoate. Octanoate growth required both lipoate protein ligase (LplA) and LipA, the sulfur insertion protein, suggesting that LplA attached octanoate to the dehydrogenase and LipA then converted the octanoate to lipoate. This pathway was tested by labeling a PDH domain with deuterated octanoate in an E. coli strain devoid of LipA activity. The labeled octanoyl domain was converted to lipoylated domain upon restoration of LipA. Moreover, octanoyl domain and octanoyl-PDH were substrates for sulfur insertion in vitro.     

Derivatives of orotic acid and its analogs

The synthesis of oxo derivatives of pyrrolo[3, 4-d]pyrimidine has been effected by the reaction of the lactone of 5-(hydroxymethyl)-orotic acid with ammonia or primary amines. In a number of cases, in place of the free organic bases their salts with mineral acids were used successfully. A number of 2, 4, 7-trioxo derivatives of pyrrolo[3, 4-d]pyrimidine and some new derivatives of orotic acid have been synthesized.     

New synthesis of pipecolic acid and analogs

Pipecolic acid congeners are synthesised from α-cyanoamides, obtained by substitution of α-methoxyamides with trimethylsilyl cyanide in an alternative route via oxidation of amidoalkylationproducts of the α-methoxyamides.     

Derivatives of orotic acid and its analogs

The lactone of 2, 6-dichloro-5-(hydroxymethyl)pyrimidine-4-carboxylic acid is synthesized, and various 2-chloro-6-amino and 2, 6-diamino derivatives are prepared from it by ammonolysis. It is shown that treatment of these compounds with alkaline reagents leads to lactone ring opening when there is a primary or secondary amino group at position 6, while the lactone ring is unaffected if there is a tertiary amino group at that position.A study is made of the action of dilute solutions of hydrochloric acid on the amino derivatives obtained by opening the lactone ring, and their reverse conversion to the corresponding 2, 6-substituted lactones of 5-(hydroxymethyl)pyrimidine-4-carboxylic acid is demonstrated.For Part III see [1].