Non-target metabolomics reveals the changes of small molecular substances in duck breast meat under different preservation time.

Poultry products are an essential animal source of protein for humans. Many factors could destroy the balance of the poultry production chain and cause an overstock of products, which need to be stored in the frozen storage warehouse for a long time. The long-term frozen storage may affect the quality of meat products. In this study, the changes of small molecular substances were revealed in duck meat during long-term storage using non-targeted metabolomics. The results showed that compared with fresh meat, even if the meat is stored under frozen storage conditions, the number of differential metabolites of frozen storage meat continues to increase with the prolongation of storage time, indicating that the meat composition has changed significantly with the storage time increased. With the increase in storage time, the nitrogen-containing small molecular compounds in duck meat increased (carnosine and anserine, aspartic acid, and tyrosine, 1H-indole-3-acetamide, 2-Hydroxyphenethylamine, 2-Naphylamine, allocystathionine, and O-phosphoethanolamine), the nucleotides decomposition process strengthened (IMP and AMP, GMP and UMP), and the content of organic acid increased (5-hydroxy indole acetic acid, 5-hydroxypentanoic acid and phenylacetate, taurine) and carbohydrate (1-O-sinapoyl-beta-d-glucose, 4-O-beta-d-glucopyranosyl-d-mannose, and alpha-d-glucose). These small molecular substances can be used as biomarkers to detect long-term stored duck meat deterioration. KEGG enrichment analysis showed that protein catabolism, nucleotide catabolism, fat decomposition and oxidation, and carbohydrate decomposition were the main metabolic processes of meat deterioration during the long-term storage of duck meat. In addition, Non-target metabolome technology is a powerful tool to reveal the meat deterioration process during long-term storage systematically. This study provided a reference for optimizing domestic poultry meat storage methods and ensuring food safety.

Metabolomic changes of the multi (-AGC-) kinase inhibitor AT13148 in cells, mice and patients are associated with NOS regulation.

INTRODUCTION: To generate biomarkers of target engagement or predictive response for multi-target drugs is challenging. One such compound is the multi-AGC kinase inhibitor AT13148. Metabolic signatures of selective signal transduction inhibitors identified in preclinical models have previously been confirmed in early clinical studies. This study explores whether metabolic signatures could be used as biomarkers for the multi-AGC kinase inhibitor AT13148. OBJECTIVES: To identify metabolomic changes of biomarkers of multi-AGC kinase inhibitor AT13148 in cells, xenograft / mouse models and in patients in a Phase I clinical study. METHODS: HILIC LC-MS/MS methods and Biocrates AbsoluteIDQ™ p180 kit were used for targeted metabolomics; followed by multivariate data analysis in SIMCA and statistical analysis in Graphpad. Metaboanalyst and String were used for network analysis. RESULTS: BT474 and PC3 cells treated with AT13148 affected metabolites which are in a gene protein metabolite network associated with Nitric oxide synthases (NOS). In mice bearing the human tumour xenografts BT474 and PC3, AT13148 treatment did not produce a common robust tumour specific metabolite change. However, AT13148 treatment of non-tumour bearing mice revealed 45 metabolites that were different from non-treated mice. These changes were also observed in patients at doses where biomarker modulation was observed. Further network analysis of these metabolites indicated enrichment for genes associated with the NOS pathway. The impact of AT13148 on the metabolite changes and the involvement of NOS-AT13148- Asymmetric dimethylarginine (ADMA) interaction were consistent with hypotension observed in patients in higher dose cohorts (160-300 mg). CONCLUSION: AT13148 affects metabolites associated with NOS in cells, mice and patients which is consistent with the clinical dose-limiting hypotension.

Development of a highly sensitive and specific monoclonal antibody based enzyme-linked immunosorbent assay for the detection of a new ß-agonist, phenylethanolamine A, in food samples.

BACKGROUND: All ß-agonists are banned as feed additives for growth promotion in animals due to toxic effects on humans after consuming the ß-agonist contaminated meats. Phenylethanolamine A (PA) is a newly emerged ß-agonist. Thus there is a need to develop highly sensitive and specific analytical methods for the detection of PA in food samples. In this study, the monoclonal antibody (mAb) against PA was produced by hybridoma technology and used for the development of enzyme-linked immunosorbent assay (ELISA). RESULTS: The IC50 values and limits of detection (LODs) of the ELISA using homogeneous combination of coating antigen/antibody for PA were 0.16 ng mL-1 and 0.011 ng mL-1 , respectively. The cross-reactive (CR) values of the assay with 14 structurally related ß-agonists were lower than 0.59%. Swine liver and meat samples were spiked with PA at different content and analysed by ELISA. Acceptable recovery rates of 91.40-105.51% and intra-assay coefficients of variation of 1.56-9.92% (n = 3) were obtained. The ELISA for seven spiked samples was confirmed by LC-MS/MS with a high correlation coefficient of 0.9881. CONCLUSION: The proposed mAb-based ELISA was highly sensitive and specific for PA and could be used as a quantitative/screening method for PA analysis in food samples. © 2016 Society of Chemical Industry.

Molecular Basis for Subtype Specificity and High-Affinity Zinc Inhibition in the GluN1-GluN2A NMDA Receptor Amino-Terminal Domain.

Zinc is vastly present in the mammalian brain and controls functions of various cell surface receptors to regulate neurotransmission. A distinctive characteristic of N-methyl-D-aspartate (NMDA) receptors containing a GluN2A subunit is that their ion channel activity is allosterically inhibited by a nano-molar concentration of zinc that binds to an extracellular domain called an amino-terminal domain (ATD). Despite physiological importance, the molecular mechanism underlying the high-affinity zinc inhibition has been incomplete because of the lack of a GluN2A ATD structure. Here we show the first crystal structures of the heterodimeric GluN1-GluN2A ATD, which provide the complete map of the high-affinity zinc-binding site and reveal distinctive features from the ATD of the GluN1-GluN2B subtype. Perturbation of hydrogen bond networks at the hinge of the GluN2A bi-lobe structure affects both zinc inhibition and open probability, supporting the general model in which the bi-lobe motion in ATD regulates the channel activity in NMDA receptors.

Subunit arrangement and phenylethanolamine binding in GluN1/GluN2B NMDA receptors.

Since it was discovered that the anti-hypertensive agent ifenprodil has neuroprotective activity through its effects on NMDA (N-methyl-D-aspartate) receptors, a determined effort has been made to understand the mechanism of action and to develop improved therapeutic compounds on the basis of this knowledge. Neurotransmission mediated by NMDA receptors is essential for basic brain development and function. These receptors form heteromeric ion channels and become activated after concurrent binding of glycine and glutamate to the GluN1 and GluN2 subunits, respectively. A functional hallmark of NMDA receptors is that their ion-channel activity is allosterically regulated by binding of small compounds to the amino-terminal domain (ATD) in a subtype-specific manner. Ifenprodil and related phenylethanolamine compounds, which specifically inhibit GluN1 and GluN2B NMDA receptors, have been intensely studied for their potential use in the treatment of various neurological disorders and diseases, including depression, Alzheimer's disease and Parkinson's disease. Despite considerable enthusiasm, mechanisms underlying the recognition of phenylethanolamines and ATD-mediated allosteric inhibition remain limited owing to a lack of structural information. Here we report that the GluN1 and GluN2B ATDs form a heterodimer and that phenylethanolamine binds at the interface between GluN1 and GluN2B, rather than within the GluN2B cleft. The crystal structure of the heterodimer formed between the GluN1b ATD from Xenopus laevis and the GluN2B ATD from Rattus norvegicus shows a highly distinct pattern of subunit arrangement that is different from the arrangements observed in homodimeric non-NMDA receptors and reveals the molecular determinants for phenylethanolamine binding. Restriction of domain movement in the bi-lobed structure of the GluN2B ATD, by engineering of an inter-subunit disulphide bond, markedly decreases sensitivity to ifenprodil, indicating that conformational freedom in the GluN2B ATD is essential for ifenprodil-mediated allosteric inhibition of NMDA receptors. These findings pave the way for improving the design of subtype-specific compounds with therapeutic value for neurological disorders and diseases.

Absence of binding of targeted analogs of somatostatin carrying cytotoxic radicals or radionuclides to growth hormone secretagogue receptors on human myocardium.

Various peripheral human tissues express receptors for growth hormone secretagogue (GHS), the highest density being in the myocardium. It was also reported that some octapeptide analogs of somatostatin (SRIH) can displace radiolabeled Tyr-Ala-hexarelin from GHS receptors on the human pituitary and heart. Thus, it is possible that radionuclide analogs of SRIH such as OctreoScan and recently developed cytotoxic SRIH analogs containing doxorubicin (DOX) intended for targeted tumor therapy, could bind to these GHS receptors, compromising the safety of compounds of this type. Therefore, we determined the binding of OctreoScan and two cytotoxic SRIH analogs consisting of octapeptide carrier RC-121 and DOX (AN-162) or 2-pyrrolino-DOX (AN-238) to human myocardium specimens. None of these compounds displayed specific binding to the human heart indicating that the clinical use of SRIH analogs linked to anthracyclines or radionuclides should not be associated with increased cardiac side effects.

[Studies on asymmetric synthesis of R-phenylethanolamine by whole cells of Arachnia sp. P163].

Effects of various factors on asymmetric synthesis of R-phenylaminoethanol from aminoacetophenone by the whole cells of Arachnia sp. P163 producing alcohol dehydrogenase for phenylethanol amine was investigated. It found that, although the reduction was inhibited by the substrate and the product, but it has the very high stereoselectivity. The reduction was normaly carried out with 2% glucose for reproduction of coenzyme in the reaction system without oxygen. The conversion yield and ee value of the product achieved 65% and 100%, respectively.

Phe310 in transmembrane VI of the alpha1B-adrenergic receptor is a key switch residue involved in activation and catecholamine ring aromatic bonding.

Pharmacophore mapping of adrenergic receptors indicates that the phenyl ring of catecholamine agonists is involved in receptor binding and activation. Here we evaluated Phe310, Phe311, and Phe303 in transmembrane VI (TMVI), as well as Tyr348 in TMVII of the alpha1B-adrenergic receptor (alpha1B-AR), which have been implicated in a catechol-ring interaction. Neither catecholamine docking studies nor mutagenesis studies of Phe311, Phe303, or Tyr348 supported a role for these residues in catechol-ring binding. By contrast, docking studies indicated that the Phe310 side chain is well positioned to interact with the catechol-ring, and substituted cysteine accessibility method studies revealed that the side chain of the 310, but not 311 residue, is both solvent accessible and directed into the agonist-binding pocket. Also, saturation mutagenesis of both Phe310 and Phe311 revealed for the former, but not for the latter, a direct relationship between side chain volume and agonist affinity, and that aromaticity is essential for wild-type agonist binding, and for both wild-type agonist potency and efficacy. Moreover, studies of Phe310 mutants combined with a previously described constitutively active alpha1B-AR mutant, A293E, indicated that although not required for spontaneous receptor isomerization from the basal state, R, to a partially activated conformation R', interaction of Phe310 with catecholamine agonists is essential for isomerization from R' to the fully activated state, R.

Phenylethanolaminotetralines compete with [3H]dihydroalprenolol binding to rat colon membranes without evidencing atypical beta-adrenergic sites.

[3H]Dihydroalprenolol ([3H]DHA) specific binding (determined by the difference in the presence and absence of 20 microM (-)isoprenaline) to rat colon membranes was saturable (Bmax = 39.6 fmol/mg protein), of high affinity (Kd = 0.87 nM) and stereospecific (IC50 330 and 3510 nM for (-)- and (+)isoprenaline, respectively); the Hill coefficient was close to one, indicating binding homogeneity. [3H]DHA (0.6 nM) specific binding was potently inhibited (Ki range 1.9-3.3 nM) by the non-selective beta-adrenoceptor antagonists pindolol, alprenolol, but not by the non-adrenergic compounds 5-hydroxytryptamine, 8-hydroxydipropylaminotetraline, methysergide, dopamine and verapamil (Ki greater than 10,000 nM). The selective beta 1- and beta 2-adrenoceptor antagonists CGP 20,712A and ICI 118,551 resulted in biphasic competition binding curves, whose low and high affinity components were compatible with two populations of binding sites accounting for about 75 (beta 2) and 25% (beta 1) of total sites. The relative competing potencies of reference adrenergic agonists also suggested a prevalence of beta 2-adrenergic sites. The new agonists phenylethanolaminotetralines (PEATs), highly selective for the atypical beta-adrenoceptors whose abundance in rat colon has been confirmed by comprehensive functional studies, had variable affinity for the [3H]DHA-labelled sites depending on chirality, but with no substantial correlation with their pharmacological potency. Only 40% of [3H]DHA binding, at a concentration about 10 times its Kd for high affinity sites (beta 1 and beta 2), was prevented by saturating concentrations of isoprenaline. Under this condition, the representative PEAT, SR 58611A, highly potent and selective for atypical beta-adrenoceptors in functional tests, and its pharmacologically inactive enantiomer, both inhibited the residual binding equipotently. In conclusion, [3H]DHA binding did not detect atypical beta-adrenoceptor sites in rat colon membranes, most probably because of its weaker affinity for them than for the coexisting beta 1 and beta 2 sites. PEAT stereoisomers proved essential for assessing both the stereospecificity and the functional significance of this atypical binding and to compare their affinity for [3H]DHA-labelled sites and pharmacological potency.

Cytochrome P-455 nm complex formation in the metabolism of phenylalkylamines. XI. Peroxygenase versus monooxygenase function of cytochrome P-450 in rat liver microsomes.

Cytochrome P-455 nm complex formation in phenobarbital induced rat liver microsomes was investigated using both an NADPH/O2-dependent monooxygenase system and a peroxygenase/peroxidase system where hydrogen peroxide was substituted for NADPH. The substrates tested were the enantiomers of four 1-alkyl-substituted 2-phenylethanamines (unbranched 1-alkyl substituents, comprising one to four carbons), S(+)- and R(-)-N-hydroxyamphetamine and racemic mixtures of N-hydroxy-1-phenyl-2-butanamine and N-hydroxy-3-methyl-1-phenyl-2-butanamine. During NADPH/O2-dependent metabolism the amines showed a positive correlation between extent of complex formation and lipophilicity; furthermore the S(+)-isomers gave rise to larger amounts of complex than the corresponding R(-)-analogues. With the hydroxylamines the ability to form complexes was greater than with any of the amines but no definite difference was seen among the hydroxylamines. In the peroxygenase system the hydroxylamines still gave larger amounts of complex than the amines but the differences seen within the homologous series of chiral amines when using the monooxygenase system were no longer observed. Although the quantitative trends in complex formation seen in the monooxygenase system were non-existent when H2O2 was substituted for NADPH, mere qualitative rules still seemed to apply; substrates which failed to give the complex during NADPH-dependent metabolism (2-phenylethanamine, phentermine, N-hydroxyphentermine and phenylacetone oxime) were inactive also in the peroxygenase system. The results substantiate the notion that the monooxygenase and peroxygenase reaction mechanisms of cyt. P-450 follow similar but not identical pathways.

Effects of aging on p- and m-octopamine, catecholamines, and their metabolizing enzymes in the rat.

Functions of octopamine in the mammalian brain are still not well known. An important aspect of this problem is the relationship between octopamines and catecholamines. Previous data have shown that their respective ontogenic evolutions are not parallel. Do the changes in brain related to aging also differentially affect these two groups of molecules? In order to check this point, the brain levels of p- and m-octopamine, p-tyramine, noradrenaline, and dopamine, as well as the activities of metabolizing enzymes, were determined in young adult and aging rats (20-26 months). Unlike catecholamines, there is a drastic decrease of p-octopamine after 20 months of age in the hypothalamus and telencephalon. p-Tyramine levels are also lowered. This change appears to be due to a decrease of the aromatic L-amino acid decarboxylase activity. These data, as those of ontogenic studies, confirm that p-octopamine and catecholamine metabolisms may have some independent steps and, moreover, that p-octopamine may have a role in the normal activity of the brain.

Prenatal ontogenesis of brain phenolamines and catecholamines in relation to their metabolizing enzymes in Roman avoider strains of rats.

Phenolamines, particularly octopamines, are of special importance in avoidance behavior. In the Roman low avoidance (RLA) strain, p-octopamine can induce locomotor behavioral activity that is normally observed in the Roman high avoidance (RHA) strain. For these reasons, the levels of prenatal octopamines (para and meta isomers) have been studied in relation to noradrenaline and dopamine levels. In the hypothalamus and brainstem of RHA, a maximum level of the para isomer is observed at 15 days of embryonic development but, unlike in controls and RLA animals, this level remains almost constant until 20 days. For the meta-isomer and catecholamines, there is a 1-2 day delay in detection between controls and RLA or RHA. The study of related enzyme activities reveals that tyrosine hydroxylase displays a 2-day delay in RHA when compared to the control value at 19 days of fetal life. These results are discussed in terms of the role of p-octopamine in avoidance conditioning and of the possible delayed expression of the tyrosine hydroxylase gene in Roman strains of rats.

The release of 3H-noradrenaline by p- and m-tyramines and -octopamines, and the effect of deuterium substitution in alpha-position.

The 3H-noradrenaline-releasing effects of p- and m-tyramines and -octopamines, either deuterated or not, were studied in isolated vasa deferentia of the rat (COMT inhibited and calcium-free solution in all experiments). Km for uptake1 was higher for octopamines than for tyramines, but not increased by the introduction of deuterium in alpha-position, except for (probably contaminated) deuterated p-octopamine. Other tissues were preloaded with 3H-noradrenaline. After inhibition of vesicular uptake and MAO equi-releasing concentrations of the eight amines were strictly correlated with Km, they were 6 to 7 times higher for unsubstituted octopamines than for corresponding tyramines. When only MAO (but not vesicular uptake) was inhibited, this difference decreased to about 4-fold, but the releasing potency of the deuterated amines (relative to their parent amines) remained unchanged (except for p-octopamine). When vesicular uptake and MAO were intact, unsubstituted octopamines were only 1.5 to 2.2 times less potent than the corresponding tyramines. Analysis of the efflux of 3H-DOPEG confirmed that this gain in the relative potencies of octopamines is due to their increased ability to mobilize vesicular 3H-noradrenaline; moreover, deuterated amines as well were then better mobilizers than were their parent amines. It is concluded that, provided vesicular uptake is intact, the introduction of a beta-OH-group enhances the ability of indirectly acting sympathomimetic amines to mobilize vesicular noradrenaline; the introduction of deuterium in alpha-position, on the other hand, enhances this mobilizing effect exclusively when MAO is intact.

Conformational and steric aspects of phenylethanolamine and phenylethylamine analogues as substrates or inhibitors of phenylethanolamine N-methyltransferase.

The conformational and steric aspects of binding to phenylethanolamine N-methyltransferase (PNMT; EC 2.1.1.28) for phenylethanolamine substrates and phenylethylamine inhibitors were probed with three conformationally defined analogues (11, 12, and 13) of phenylethylamine (1) and phenylethanolamine (6) containing the benzobicyclo[3.2.1]octane skeleton. The 2-aminotetralin (2AT) moiety in conformationally defined analogues 11, 12, and 13 exists in a half-chair conformation with an equatorial amino group. Although conformationally restricted phenylethylamine analogue 2AT (3, Ki = 6.8 microM) and conformationally restricted phenylethanolamine analogues (cis)- and (trans)-2-amino-1-tetralol (9, Km = 22 microM; Vmax = 0.15; 100 X Vmax/Km = 0.68; 10, Ki = 9.4 microM) are good ligands for PNMT, none of the analogues 11, 12, and 13 showed activity as a substrate of PNMT. The fact that 11 (Ki = 206 microM) is more potent than analogues 4 (Ki = 1296 microM) and 5 (Ki = 479 microM), with a half-boat 2AT moiety, suggests that PNMT preferentially binds the half-chair conformation of 2AT at the active site. This is consistent with previous findings that a fully extended conformation for the aminoethyl side chain of phenylethylamine inhibitors is optimal for PNMT binding. The reduced activity of 11, 12 (Ki = 1246 microM), and 13 (Ki = 3000 microM), compared with 2AT and (cis)- and (trans)-2-amino-1-tetralol (9 and 10) is consistent with a negative steric interference from the extra ethano bridge in 11, 12, and 13. The results from 11, 12, and 13, combined with previous findings, suggest that PNMT interacts better with relatively planar ligands.

Biochemical studies of N-methyltransferase in human and guinea-pig lung: no apparent role in the pathogenesis of asthma.

1. N-Methyltransferase activity was measured in surgical specimens of human lung using phenylethanolamine as substrate. Thirty-three male and seven female patients, age range 19-78 (median 62.5) years were studied. The activity in lung homogenates was 0.59 x 10(-6) units/mg of protein (SEM 0.03, n = 40), with a range of 0.16-1.16 x 10(-6) units/mg of protein. There was no difference (P = 0.97) in activity between males and females. 2. Non-specific N-methyltransferase activity was estimated in 17 of the surgical specimens using beta-phenylethylamine as substrate. This activity was 38.9% (SEM 5.3) of that with phenylethanolamine. Comparative studies with rabbit lung, which has a well-characterized non-specific N-methyltransferase, showed significant differences in substrate specificity between the two species. 3. The apparent Km and Vmax for phenylethanolamine in seven human lung homogenates was 22.0 (SEM 4.6).mmol/l and 1.82 x 10(-6) units/mg of protein (SEM 0.36). The noradrenaline N-methyltransferase (NMT; EC 2.1.1.28) inhibitors SKF 64139-A and LY 134046 did not inhibit this activity up to a concentration of 100 mumol/l. This activity was inhibited 51.4% (SEM 8.6, n = 6) by 100 mumol/l S-adenosyl-L-homocysteine. Immunohistochemistry did not reveal immunoreactive NMT in human lung sections. 4. Comparative studies with guinea-pig lung homogenates demonstrated non-specific N-methyltransferase activity in this species which is similar to the human lung.(ABSTRACT TRUNCATED AT 250 WORDS)

Activities of octopamine and synephrine stereoisomers on alpha-adrenoceptors.

1. The activities of the (-)- and (+)-forms of m- and p-octopamine and m- and p-synephrine on alpha 1-adrenoceptors from rat aorta and anococcygeus and alpha 2-adrenoceptors from rabbit saphenous vein were compared with those of noradrenaline (NA). 2. The rank order of potency of the (-)-forms on alpha 1-adrenoceptors from rat aorta and alpha 2-adrenoceptors was NA greater than m-octopamine = m-synephrine greater than p-octopamine = p-synephrine. The two m-compounds were 6 fold less active than NA on alpha 1-adrenoceptors from rat aorta and 150 fold less active on alpha 2-adrenoceptors. The two p- compounds were 1,000 fold less active than NA on both alpha 1-adrenoceptors from rat aorta and alpha 2-adrenoceptors. The rank order of potency of the (-)- forms on alpha 1-adrenoceptors from rat anococcygeus was NA = m-synephrine greater than m-octopamine greater than p-octopamine = p-synephrine. m-Octopamine was 4 fold less active than NA and (-)-m-synephrine. The two p- compounds were 30 fold less active than NA. 3. The rank order of potency of the (+)- forms was NA greater than m-octopamine greater than m-synephrine greater than p-octopamine greater than p-synephrine on both alpha 1- and alpha 2-adrenoceptors. The potency of each (+)- form was 1-2 orders of magnitude less than that of the (-) counterpart, the differences being greater for the stereoisomers of synephrine than for those of octopamine on both alpha 1- and alpha 2-adrenoceptors. 4. The yohimbine diastereoisomer antagonists, rauwolscine and corynanthine, were tested against (-)-NA and (-)-m-octopamine-induced contractions in both preparations. Based upon the known selectivities of these isomers for alpha-adrenoceptor subtypes, it is concluded that the rat aorta contains only alpha 1-adrenoceptors while the rabbit saphenous vein possesses predominantly alpha 2-adrenoceptors. 5. Ligand binding data for the octopamine and synephrine stereoisomers at alpha 1- and alpha 2-binding sites from rat cerebral cortex was also obtained. (-)-Forms were more active than (+)-forms. The rank order of affinity of the (-)-forms for both alpha 1- and alpha 2-binding sites was NA greater than m-octopamine = m-synephrine greater than p-synephrine greater than p-octopamine. The relative affinities of the members of the series against alpha 1-binding sites were very similar to their relative functional activities on rat aorta. However, the affinities of both m- and p-compounds relative to that of ( -)-NA were much greater at the x2-binding sites than were the relative activities in rabbit saphenous vein, possibly suggesting low intrinsic efficacy. Functional antagonist responses to NA by the (-)-octopamine and synephrines could not, however, be demonstrated on rat aorta or rabbit saphenous vein. 6. The activities of m-octopamine and m-synephrine were not significantly different from each other on either a,-adrenoceptors from rat aorta or x2-adrenoceptors; however, m-synephrine is more active than m-octopamine on a,-adrenoceptors from rat anococcygeus. Both m-octopamine and msynephrine can be considered to be naturally occurring x,-selective amines. However, if m- and poctopamine are co-released with NA in amounts proportional to their concentration, it is concluded that their activities on m,- and x2-adrenoceptors are too low to be physiologically significant.

Excretion of octopamine metabolites in neuroblastoma.

The urinary concentrations of o-hydroxymandelic acid, m-hydroxymandelic acid, p-hydroxymandelic acid, homovanillic acid and vanillylmandelic acid were determined in 57 healthy children and 9 patients with neuroblastoma. The concentrations of o-hydroxymandelic acid and p-hydroxymandelic were not significantly different for both groups whereas the concentrations of m-hydroxymandelic acid, homovanillic acid and vanillylmandelic acid were elevated 20- to 30-fold in the neuroblastoma patients.

The concentration in brain of octopamine and tyramine after portal-systemic bypass in rats: neuroamine concentrations determined simultaneously by methane chemical ionization gas chromatography mass spectrometry.

The concentration in brain of both octopamine (OCT) and tyramine (TYR) was significantly increased in rats 8 weeks after portal-systemic bypass. This suggests that the increase in OCT is secondary to increased decarboxylation of tyrosine to TYR. However, the role these neuroamines, particularly OCT, play in the development of hepatic encephalopathy remains controversial.

High performance liquid chromatography coupled with radioactivity detection: a powerful tool for determining drug metabolite profiles in biological fluids.

High performance liquid chromatography coupled with continuous radioactivity detection represents an advancement in drug metabolism research. Using radioactive substances labelled in biologically stable positions, all metabolites can be specifically detected by radioactivity measurement. Thus no clean-up of biological fluids is required prior to HPLC. This can prevent artefact formation from unstable metabolites, reduces recovery problems and facilitates quantitation. Separation of highly polar and unpolar metabolites is possible in a single chromatographic run using gradient elution and reversed phase materials. This technique is also well-suited for preparative isolation and purification of metabolites for subsequent structure elucidation. Various metabolite profiles of drugs labelled with carbon-14 or tritium are shown. Metabolites of the following drugs are presented: norfenefrine, etozolin, thymoxamine, naloxone, and levobunolol. We review the general methodology and report our experience with this technique. In principle, this technique may be useful for all biological systems in which tracer techniques are applied.

The brain octopamine and phenylethanolamine content in rats in thioacetamide-induced hepatogenic encephalopathy.

The brain octopamine (OA) and phenylethanolamine (PhEA) content was determined in rats subjected to repeated intraperitoneal administrations of thioacetamide (TAA) known to produce different stages of hepatogenic encephalopathy (HE). A more than 2.5-fold increase of OA and a 2-fold increase of PhEA was observed after prolonged (3 times) administration of TAA, coinciding with impaired ammonia detoxication in brain and with the onset of pathophysiological changes typical for HE. Only insignificant changes in the content of the amines were observed in the early stages of the experiment as well as in the recovery period. The results are consistent with the "false neurotransmitter" hypothesis of Fischer and Baldessarini, assuming the participation of OA and PhEA in the pathogenesis of HE.