Δ-aminovaleric Acid Research Articles

Inhibitors for the in vitro assembly of Lp(a).

Lp(a) is composed of an LDL-like core and the glycoprotein apo(a). Current evidence strongly suggests that the assembly of this atherogenic lipoprotein proceeds outside the liver cells in a two-step fashion. In the first step, a loose complex is formed involving kringle-4 motifs in apo(a) and one or more Lys side chains in apoB-100. In the second step, this complex is stabilized by a disulfide bridge. Indications are that Lp(a) assembly is critical in the determination of plasma apo(a) concentrations. Therefore, we searched for substances that interfere with the first step of Lp(a) assembly. epsilon-Aminohexoic acid (epsilon-AHA), known as an inhibitor from earlier assembly studies, had an IC50 of 4.8 mmol/L. The IC50 of Pro, HO-p-aminobenzene sulfonamide, Lys, N-epsilon-acetyl-Lys, taurine, Glu, serotonin, and benzamidine were all > 20 mmol/L. gamma-Aminobutyric acid, spermine, and spermidine exhibited IC50 on the same order of magnitude as epsilon-AHA. The substances with the highest inhibitory action were tranexamic acid and delta-aminovaleric acid. Seven of eight patients treated in a pilot study with tranexamic acid (Cyclocapron) responded with a decrease of plasma apo(a) of 18.5 +/- 8.2%. We suggest that substances that interfere with the Lp(a) assembly are worth pursuing further for their usefulness as therapeutic agents in reducing high plasma Lp(a) concentrations.

Functional role of GABA in cat retina: I. Effects of GABAA agonists.

Putative GABAergic mechanisms were studied in perfused cat retina by means of intracellular recording and application of GABA and the GABAA agonists delta-amino valeric acid (dAVA), muscimol, and THIP. In contrast to results reported previously for cold-blooded vertebrates, introduction of 20 mM GABA into the superfusate had no influence upon the response properties of cat retinal horizontal cells (HCs). In common with results reported in cold-blooded vertebrates, introduction of the GABAA agonists dAVA (2-12 mM) and THIP or muscimol (0.2-1 mM) had four consistent reversible influences upon the response properties of cat retinal HCs: (1) they reduced photic-response amplitude, (2) slowed response onset, (3) slowed response offset, and (4) depolarized the dark membrane potential. Both rod and cone signal components were affected. GABAA agonists had similar influences upon both the time course and amplitude of responses recorded from amacrine and ganglion cells. In all cell types examined, the influence upon response kinetics was made particularly apparent with rapidly flickering stimuli. Flicker responses were reduced in amplitude much more than sustained responses. These results suggest that, in addition to other influences, GABAergic action serves to modify the time course of photic responses in both the inner and outer plexiform layer of mammalian retina making responses slower and less phasic.

Quantitative estimation of betaines in commercial seaweed extracts using overpressured layer chromatography

An off-line, overpressured layer chromatographic procedure has been developed for the evaluation of betaines in commercial seaweed extracts using Dragendorff's reagent for the detection of the compounds and densitometry for their quantitative estimation. Using continuous development and a low sample volume, the amounts of γ-aminobutyric acid betaine and δ-aminovaleric acid betaine can be estimated, but a larger sample volume is necessary for the estimation of glycinebetaine and minor betaines. The critical steps of this analytical method are the visualization and the quantitative evaluation of the spots produced. Temperature plays a major role in the resolution, sensitivity and precision after derivatization with Dragendorff's reagent; this has led to the adoption of standardized conditions.

The preparation and characterization of some Amadori compounds (1-amino-1-deoxy- d-fructose derivatives) derived from a series of aliphatic ω-amino acids

Amadori compounds (1-amino-1-deoxy- d-fructose derivatives) were prepared by reacting d-glucose with a series of aliphatic amino acids. These include Amadori compounds derived from glycine ( 1), β-alanine ( 2), γ-amino butyric acid ( 3), δ-aminovaleric acid ( 4), ϵ-aminocaproic acid ( 5) and N α-formyl- l-lysine ( 6). In the FAB mass spectra, molecular-ion clusters as well as fragment ions corresponding to loss of water or CO 2 molecules were observed. The 13C NMR spectra indicate that all the compounds are conformationally unstable, but that the predominant from present in solution (D 2O) is the β-pyranose form. The 1H NMR spectra of 1 and 2 indicate a slow rotation around the C-1—C-2 bond, possibly as a result of an intramolecular hydrogen bond involving the carboxyl group. The p K a 's of all compounds were measured by pH-potentiometric titration in 0.2 M KNO 3 solution at 25°C. All compounds showed a decrease in the basicity of their amino groups (in the order of ≈ 1.5 of the K a value), and 1 and 2 showed a decrease in the basicity of their carboxyl groups (in the order of ≈ 0.2) in comparison with that of parent amino acids.

Analysis of hydrogen bonds in peptides, based on the hydration affinity of amides

Abstract The difference in the affinity for water of peptide groups embedded in different molecular environments was investigated. The chemical shift of an amide proton is sensitive to conformational variations, as well as to changes in the molecular environment [D.S. Wishat, B.D. Sykes and F.M. Richards, J. Mol. Biol., 222 (1991) 311–333]. Therefore, if the conformational motions are minimized or excluded, the observed changes in the chemical shift can simply be related to the environmental effects. The conformation(s) of the cyclic β-turn models studied in this work has been previously reported using X-ray, NMR, circular dichroism, and (FT-IR) spectroscopic methods, as well as MD calculations. [M. Hollosi, K.E. Kover, S. Holly, L. Radics and G.D. Fasman, Biopolymers, 26 (1987) 1527–1572; A. Perczel, M. Hollosi, B.M. Foxman and G.D. Fasman, J. Am. Chem. Soc., 113 (1991) 9772–9784; and H.H. Mantsch, A. Perczel, M. Hollosi and G.D. Fasman, Biopolymers, 33 (1993) 201–207]. The backbone of the cyclo[(δ)Ava—Gly—Pro—Aaa—Gly] (where Aaa = Ser(O t Bu), Ser or Thr(O t Bu), and δ(Ava) is δ-aminovaleric acid) compounds was found to be rigidly incorporated in the structure and to contain two intramolecular hydrogen bonds. These β-turn models also include one (or two) “free” amide group(s) that are not involved in any type of interaction. The “water titration” of these amide groups in acetonitrile, where they are involved in various degrees of hydrogen bonding, revealed their molecular environment. Owing to the rigidity of these structures, the observed changes in the amide proton chemical shifts, during titration were attributed to their involvement in hydrogen bonding. This was confirmed by monitoring the water titration simultaneously with FT-IR spectroscopy. The phenomenon described here, with the proposed characterization of the investigated peptide/water system, comprise an improvement in the NMR method for analyzing the hydrogen bonding of small rigid peptides.

The Action of Diazepam in the Isolated Rat Detrusor Muscle

Diazepam is one of the benzodiazepines, a group of drugs that depresses the central nervous system. It also inhibits the contractility of smooth muscles in the periphery, but the mechanism of this inhibitory action has not been clarified. Our study was undertaken to investigate the effect of diazepam on the contractility of the detrusor muscle. Detrusor muscle strips isolated from rat urinary bladder were examined by isometric myography.Diazepam, as well as baclofen, a γ-aminobutyric acid (GABA)B receptor agonist, reduced the electric field stimulation-induced contractions; δ-aminovaleric acid, a GABAB receptor antagonist, completely antagonized the inhibitory effect of baclofen, but not the inhibitory action of diazepam. Diazepam reduced the basal tone of detrusor muscle concentration dependently, and this inhibitory action was not affected by tetrodotoxin. Diazepam suppressed the contractile responses to betha-nechol, adenosine triphosphate and potassium chloride. Diazepam diminished the calcium-induced recovery of tension in calcium-free PSS. A23187, a calcium ionophore, partially recovered the basal tone which had been reduced by diazepam in normal physiologic salt solution (PSS). The loss of tension in calcium free PSS containing diazepam could not be recovered by addition of A23187. On the other hand, the loss of tension in calcium-free PSS containing 3,4,5-trimethoxybenzoic acid 8(diethylamino)octyl ester (TMB-8), an inhibitor of intracellular calcium release, was considerably recovered by addition of A23187.Based on these results, it is suggested that diazepam inhibits the contractility of detrusor muscle acting directly on the smooth muscle cell, which is unrelated to the activation of GABA receptors. Its inhibitory action appears to be mediated through interference with the influx of extracellular calcium.

Significance of betaines in the increased chlorophyll content of plants treated with seaweed extract

Seaweed extract, prepared by alkaline extraction of Ascophyllum nodosum (L.) Le Jol., applied either to the soil or to the foliage of tomato plants, produced leaves with higher chlorophyll levels than those of control plants. The effects on leaf chlorophyll content were investigated using a cucumber bioassay procedure devised for cytokinins. The seaweed extract was shown to increase the chlorophyll levels of the cucumber cotyledons, but ‘peaks’ of activity were obtained when widely different concentrations were used. The possibility that these effects were the result of betaines present in the extract was considered. Glycinebetaine, γ-aminobutyric acid betaine and δ-aminovaleric acid betaine all produced significantly enhanced chlorophyll concentrations in the cotyledons. ‘Peaks’ of activity were observed for each betaine: for glycinebetaine at 10−6 and between 10−4 and 101 mg 1−1, for γ-aminobutyric acid betaine at 10−6, between 10−4 and 10−1, and 101 mg 1−1, and for δ-aminovaleric acid betaine between 10−5 and 101 mg 1−1. It was concluded that the effects of enhancing chlorophyll levels produced by the seaweed extract were due, at least in part, to betaines.

Betaines and tertiary sulphonium compounds from 62 species of marine algae

From all 62 species of marine algae studied, either betaines or a tertiary sulphonium analogue or a mixture of the two types of compounds were extracted. Thus from this and earlier studies it would appear that either one or both of these classes of compounds are a constant feature of marine algae. Some compounds, in particular glycinebetaine, γ-aminobutyric acid betaine, prolinebetaine and 3-dimethyl-sulphoniopropionate, have a common and widespread distribution, whereas other compounds, e.g. β-prolinebetaine, trans-4- hydroxy-β-prolinebetaine , N- acetyllamine , δ-aminovaleric acid betaine and α-alaninebetaine, appear to have a very restricted occurrence. From Ptilota serrata, γ-aminobutyric acid betaine, methyl ester, was isolated and characterized; this is the first record of this substance as a natural product. It was not detected in any of the other algae investigated. The yields of betaines and tertiary sulphonium compounds obtained by extraction of the algae were low, especially from the Phaeophyceae. Apart from Chaetomorpha capillaris, for which 2% (of dry weight) of glycinebetaine was recorded, the yield of the major betaine or tertiary sulphonium compound from the other species investigated was less than 1% (of dry weight) and frequently less than 0.1%. The similarity of the betaines and their tertiary sulphonium analogues found in different species of the same genus show that these compounds have taxonomic significance at the genetic level.

3-aminopropanephosphinic acid is a potent agonist at peripheral and central presynaptic GABA B receptors

The actions of the GABA analog 3-aminopropanephosphinic acid (3-APA) were studied in the guinea-pig isolated ileal preparation and at synapses between cultured rat hippocampal neurons. Like the GABA B receptor agonist, baclofen, 3-APA inhibited the electrically evoked ileal twitch. The EC 50 for 3-APA was 0.8μM; the EC 50 for baclofen was 9 μM. In addition, the depressant responses to 3-APA and baclofen were blocked by the GABA B receptor antagonists phaclofen, saclofen, 2-hydroxy-saclofen and δ-aminovaleric acid. 3-APA also mimicked the presynaptic action of baclofen at GABAergic synapses between embryonic rat hippocampal neurons in culture. 3-APA reduced the amplitude of inhibitory postsynaptic potentials (IPSPs) and currents (IPSCs) by greater than 50% at a concentration of 1 μM, while baclofen reduced synaptic transmission to a similar degree at 10 μM. 3-APA did not alter membrane conductance, nor did the drug alter postsynaptic responses to GABA. These data show that 3-APA is a potent agonist at presynaptic GABA B receptors in the periphery and on GABAergic neurons from the central nervous system. The activity of 3-APA at central postsynaptic GABA B receptors remains to be studied.

Some characteristics of baclofen-evoked responses of primary afferents in frog spinal cord

Baclofen has been shown to be a selective agonist for a subclass of GABA receptors (GABA B) in many regions of the vertebrate nervous system. On the intraspinal terminals of dorsal roots (DRT), it evokes a pure hyperpolarizing response. We have previously shown that the response of DRT to GABA and some of its analogs (e.g. kojic amine) in isolated frog spinal cord is dual in nature, consisting of a bicuculline-sensitive depolarizing component and a bicuculline-resistant hyperpolarizing component. Under the working hypothesis that the hyperpolarizing component of the GABA-evoked response is mediated by the activation of GABA B receptors, we have examined, using the sucrose gap technique, some characteristics of the response of DRT to baclofen. We have found that this response is stereospecific ( l-baclofen being about 100 times more potent than d-baclofen), dependent on [K] o (response amplitude inversely related to [K] o), blocked by barium (0.5 mM causing a reduction of the response amplitude to 37% of control), and is not significantly affected by 4-aminopyridine, nor by inorganic calcium channel blockers (manganese, cobalt, cadmium). Some proposed GABA B antagonists (δ-aminovaleric acid, δ-aminolaevulinic acid, phaclofen) are also rather ineffective at blocking it. These results are therefore consistent with the notion that the baclofen-evoked response of DRT is mediated by an increase in conductance to potassium ions.

Free amino acid levels in oral fluids of normal subjects and patients with periodontal disease

The content of free amino acids in submandibular, parotid and whole saliva and in gingival fluid was measured by automatic analyser in patients with progressive periodontitis and in controls. The concentration of individual amino acids showed considerable variation in all the fluids. Two amino acids, proline and δ-aminovaleric acid, were never found in gingival fluid or in submandibular and parotid saliva, although detectable in whole saliva. Thus, these amino acids must appear in whole saliva as a result of bacterial metabolism or by degradation of salivary proteins rich in proline. The free amino acid levels in whole saliva did not reflect those of any of the single oral fluids analysed, further stressing the importance of bacterial metabolism as a cause of the wide range of amino acid concentrations. Because of the multifactorial origin of these free amino acids, their determination in oral fluids is of no practical diagnostic value except perhaps in extreme cases.

The effects of GABA B receptor agonists and antagonists on potassium-stimulated [Ca 2+] i in rat brain synaptosomes

The effects of GABA B agonists and putative antagonists on intrasynaptosomal calcium ion concentrations ([Ca 2+] i) after stimulation with potassium ions were studied with the fluorescent probe Quin 2. γ-Aminobutyric acid and (−)-baclofen, but not (+)-baclofen, produced a dose-dependent inhibition of the potassium-stimulated [Ca 2+] i in cortical synaptosomes from the rat. This effect was mimicked by another GABA B agonist SL75102 and weakly by muscimol. It was not inhibited by the α-adrenoceptor antagonist phentolamine. This system thus appears to provide a useful test of GABA B receptor function. None of the putative GABA B antagonists, phaclofen, δ-amino valeric acid or β-phenyl GABA inhibited responses to (−)-baclofen. Indeed, all three compounds produced similar responses to that seen with (−)-baclofen, suggesting that they act as agonists in this system. These data suggest that those GABA B receptors modulating [Ca 2+] i have a distinct pharmacology from post-synaptic GABA B receptors, defined in electrophysiological experiments.

GABAB receptor antagonist and GABAA receptor agonist properties of a delta-aminovaleric acid derivative, Z-5-aminopent-2-enoic acid.

The activity of Z-5-aminopent-2-enoic acid (A), a conformationally restricted analogue of delta-aminovaleric acid, was investigated in vitro in the guinea pig isolated ileum and vas deferens. A was found to be a relatively potent GABAB-receptor antagonist and a relatively weak GABAA-receptor agonist. The GABAB-receptor antagonist activity of A was at least 5 times greater than that of phaclofen, a selective GABAB-receptor antagonist. A may therefore be an important lead for the development of new potent and selective GABAB-antagonists.

ChemInform Abstract: New Heterobifunctional Cross‐Linking Reagents for Protein Conjugation, N‐(Bromoacetamido‐n‐alkanoyloxy)succinimides.

Five heterobifunctional reagents, N- (bromoacetamido-n-alkanoyloxy) succinimides having a glycine or ω-amino acid (β-alanine, γ-aminobutyric acid, δ-aminovaleric acid or e-aminocaproic acid) residue, were synthesized. Bromoacetamido-n-alkanoyl groups could be introduced into horseradish peroxidase and hen egg-white lysozyme and its reduced and S-3- (trimethylated amino) propylated product by using the reagents. The number of bromoacetamido-n-alkanoyl groups introduced into the proteins could be successfully estimated from the amount of glycine or ω-amino acid (non-protein amino acid) produced from the introduced groups by acid hydrolysis. N- (β-Bromoacetamido-n-propionoyloxy) succinimide, one of the reagents, was examined as a reagent for the preparation of horseradish peroxidase-insulin conjugate. A single β-bromoacet- amido-n-propionoyl group was first introduced into GlyA1, PheB1-dicitraconylinsulin through the e-amino group of its LysineB29 residue and the product was subjected to decitraconylation with dilute acetic acid to obtain LysB29- (β-bromoacetamido-n-propionoyl) insulin. The insulin was reacted with thiolated horseradish peroxidase to give peroxidase-insulin conjugate (molar ratio of 1 : 1).

Differing actions of β-phenyl-GABA and baclofen in the guinea pig isolated ileum

In the guinea-pig isolated ileum, both γ-aminobutyric acid (GABA) and baclofen induced a dose-dependent depression of cholinergic twitch contractions to transmural stimulation, sensitive to δ-aminovaleric acid (DAVA) and phosphonobaclofen (phaclofen). β-Phenyl-GABA (BPG) antagonised this depressant action of baclofen and GABA, whilst itself weakly depressing ileal twitch contractions, an effect insensitive to DAVA or phaclofen, and thus unrelated to any GABA B-receptor-mediated effects. These results suggest that the baclofen receptors in the ileum that are antagonised by BPG differ from either of the baclofen receptors in the spinal cord, where the presynaptic receptors are blocked by phaclofen and the postsynaptic receptors are insensitive to phaclofen, with BPG having baclofen-like actions at both sites. Interaction of BPG and baclofen with different receptor populations may explain the differing therapeutic actions of these compounds.

Comparison of GABA-induced responses in various segments of the guinea-pig intestine

In preloaded, isolated segments of duodenum, jejunum, ileum and colon, ethylenediamine (EDA) caused a 3-mercaptopropionic acid (3-MPA)-sensitive release of [ 3H]GABA. Both EDA and GABA caused a contraction and a delayed ‘after-relaxation’ in the ileum, but only a δ-aminovaleric acid (DAVA)-sensitive relaxation in other intestinal segments; these actions of EDA were reduced by 3-MPA and therefore due to release of endogenous GABA. Baclofen induced a DAVA-sensitive relaxation in all tissues. Evidently, GABA modulates cholinergic excitation of the smooth muscle at all levels of the intestine.

Free amino-acid content of wax-stimulated human whole saliva as related to periodontal disease

This content was analysed in patients with chronic periodontitis and in control subjects. In periodontal disease, it was characterized by higher mean concentrations of glycine, proline, tyrosine and (δ-aminovaleric acid than in controls ( p < 0.001). However, the range of values varied considerably in the two groups. There were differences between periodontitis and control samples in the proportions of proline to serine ( p < 0.01) and proline to glutamic acid and glutamine ( p < 0.05). Bacterial contamination and decomposition of salivary proteins is responsible for the elevated salivary levels of glycine, proline, tyrosine and δ-aminovaleric acid in the periodontal group.

New heterobifunctional cross-linking reagents for protein conjugation, N-(bromoacetamido-n-alkanoyloxy)succinimides.

Five heterobifunctional reagents, N- (bromoacetamido-n-alkanoyloxy) succinimides having a glycine or ω-amino acid (β-alanine, γ-aminobutyric acid, δ-aminovaleric acid or e-aminocaproic acid) residue, were synthesized. Bromoacetamido-n-alkanoyl groups could be introduced into horseradish peroxidase and hen egg-white lysozyme and its reduced and S-3- (trimethylated amino) propylated product by using the reagents. The number of bromoacetamido-n-alkanoyl groups introduced into the proteins could be successfully estimated from the amount of glycine or ω-amino acid (non-protein amino acid) produced from the introduced groups by acid hydrolysis. N- (β-Bromoacetamido-n-propionoyloxy) succinimide, one of the reagents, was examined as a reagent for the preparation of horseradish peroxidase-insulin conjugate. A single β-bromoacet- amido-n-propionoyl group was first introduced into GlyA1, PheB1-dicitraconylinsulin through the e-amino group of its LysineB29 residue and the product was subjected to decitraconylation with dilute acetic acid to obtain LysB29- (β-bromoacetamido-n-propionoyl) insulin. The insulin was reacted with thiolated horseradish peroxidase to give peroxidase-insulin conjugate (molar ratio of 1 : 1).

31] Synthesis of carnitine precursors and related compounds

This chapter discusses the synthesis of carnitine precursors and related compounds. L-Carnitine contains a quaternary nitrogen atom which is formed biosynthetically early in the pathway. Thus several intermediates in the pathway are quaternary amines which can readily be synthesized from the commercially available parent primary amines. These intermediates include ɛ-N-trimethyl-L-lysine (from α-N-acetyl-L-lysine), γ-N-trimethylaminobutyraldehyde (from 4-aminobutyraldehyde diethyl acetal), and γ-butyrobetaine (from γ- aminobutyric acid). Other compounds of interest which are synthesized by this general method include δ- N -trimethylaminovaleric acid (from δ-aminovaleric acid) and ɛ-N-trimethylaminocaproic acid (from ɛ-aminocaproic acid). Another carnitine precursor, β -hydroxy- ɛ-N- trimethyl-L-lysine, is prepared by condensation of γ- N -trimethylaminobutyraldehyde with glycine in the presence of potassium carbonate and copper sulfate, yielding a diastereomeric mixture. Carnitine is synthesized, as a racemic mixture, by several related methods. For example, DL-carnitine is prepared by reaction of epichlorhydrin with NaCN followed by condensation with trimethylamine, and subsequent hydrolysis. Neurospora crassa (but apparently not in mammals) ɛ- N -methyl-L-lysine and ɛ- N -dimethyl-L-lysine are direct precursors of L-carnitine. The latter compound is prepared by reductive methylation of α-N-acetyl- L-lysine with formaldehyde and H 2 /Pd catalyst. α-Keto-ɛ- N -trimethylaminocaproic acid, a catabolic product of ɛ-N-trimethyl- L-lysine, is synthesized enzymatically by reaction of ɛ- N -trimethyl- L-lysine with commercial L-amino acid oxidase. The chapter describes the representative synthetic procedures for ɛ- N - Trimethyl-L-lysine, ɛ- N -[methyl- 14 C]DimethyI-L-lysine, β -Hydroxy-ɛ- N -[methyl- 3 H]trimethyl-L-lysine, γ-[1- 14 C]Butyrobetaine, and L-[methyl- 3 H] Carnitine.

Identification of extrasynaptic binding sites for [ 3H]GABA in peripheral nerve

The binding of radiolabelled γ-aminobutyric acid ([ 3H]GABA) to extrasynaptic sites in peripheral nerve was examined in order to characterize the receptor species mediating the depolarizing action of GABA on myelinated axons. Binding of [ 3H]GABA was carried out under Na +-free conditions with fresh homogenates of amphibian sciatic nerve. The kinetics of association of the radioligand suggested the presence of a rapidly associating, reversible binding site, and a slowly associating, apparently irreversible one. Scatchard plots of the reversible component of binding were linear and yielded a mean affinity (K d) of 22 nM. This stands in contrast with the ED 50 of 90 μM for GABA-evoked depolarization of frog nerve; and therefore the high affinity binding sites are unlikely to be involved in such depolarization. The rank order of potency of agonists that competedwith [ 3H]GABA was muscimol > GABA≫ δ-aminovaleric acid> β-alanine. However, the GABA A analogues 3-aminopropanesulfonic acid, β-guanidinopropionic acid, and guanidoacetic acid as well as the GABA B agonist baclofen all failed to displace the ligand. Bicuculline methiodide, picrotoxin, and nipecotic acid also did not compete. The bicuculline-insensitive, baclofen-insensitive high affinity binding sites identified here appear to be unique, as they are distinctly different from the classical GABA A and GABA B receptors.