RESUMEN
Serotonin (5-HT), its precursor 5-hydroxytryptophan (5-HTP), and its major metabolite 5-hydroxyindoleacetic acid (5-HIAA) were measured in the cerebrospinal fluid (CSF) of 14 patients with dementia of the Alzheimer type (DAT) and in nine controls by high-performance liquid chromatography with a novel multisensor coulometric detection system. Concentrations of both 5-HT and 5-HIAA detected by this system were lower than the concentrations obtained using conventional amperometric detection. This difference was caused by coelution of compounds that could be resolved from 5-HT and 5-HIAA by the multisensor coulometric system. One of the coelution compounds, observed in DAT but not in control CSF, behaved like a partially oxidized 5-HT. A compound behaving like partially oxidized 5-HTP was also observed in DAT CSF. Concentrations of 5-HTP, 5-HT, and 5-HIAA were lower in DAT CSF than in a corresponding fraction of control CSF. These results indicate involvement of the serotoninergic system in DAT and might lead to development of a diagnostic test for DAT.
Asunto(s)
5-Hidroxitriptófano/líquido cefalorraquídeo , Enfermedad de Alzheimer/líquido cefalorraquídeo , Demencia/líquido cefalorraquídeo , Ácido Hidroxiindolacético/líquido cefalorraquídeo , Anciano , Humanos , Persona de Mediana EdadRESUMEN
It has been demonstrated that 5-hydroxytryptamine (5-HT) is not the only neuroactive metabolite of tryptophan (TRP) in the CNS. The presence of kynurenine (KYN) and its metabolites has been reported in the brain of several mammalian species and the neuroactive properties of these compounds are now well established. In the present study, we report the identification of KYN in the superficial layers of the rat spinal dorsal horn. KYN was measured simultaneously with TRP. 5-hydroxytryptophan, 5-HT, 5-hydroxyindoleacetic acid and 5-HT-O-sulfate by means of liquid chromatography with coulometric electrode array detection. The results observed in the normal rat and in an animal model of persistent pain, the arthritic rat, are discussed in view of the hypothesis relating to the involvement of the bulbospinal serotonergic system in pain mechanisms and of the possible participation of KYN and its metabolites in these mechanisms.
Asunto(s)
Artritis/metabolismo , Quinurenina/análisis , Serotonina/análisis , Médula Espinal/metabolismo , Triptófano/metabolismo , Animales , Región Lumbosacra , Ratas , Valores de Referencia , Triptófano/análisisRESUMEN
Six female cats, varying in susceptibility to motion sickness, were implanted with chronic cannulae in the rostral portion of the fourth ventricle. The cats were then challenged with a motion sickness-inducing stimulus. Samples of cerebrospinal fluid were withdrawn before and after emesis or 30 min of motion if emesis did not occur and again on control (no motion) days. The samples were analyzed by HPLC with an array of 16 coulometric detectors. Thirty-six compounds were identified in the samples. Baseline levels of DOPAC, MHPGSO4, uric acid, DA, 5-HIAA and HVA were lower on motion and control days in cats which became motion sick when compared with cats which did not become motion sick. None of the identified compounds varied as a function of either exposure to motion or provocation of emesis. It is concluded that susceptibility to motion sickness is a manifestation of individual differences related to fundamental neurochemical composition.
Asunto(s)
Gatos/líquido cefalorraquídeo , Mareo por Movimiento/líquido cefalorraquídeo , Animales , Susceptibilidad a Enfermedades , Dopamina/líquido cefalorraquídeo , Femenino , Ácido Homovanílico/líquido cefalorraquídeo , Ácido Hidroxiindolacético/líquido cefalorraquídeo , Mareo por Movimiento/fisiopatología , VómitosRESUMEN
Recent hypotheses and findings indicate that measurements of interactions between cerebrospinal fluid (CSF) biogenic amine systems, rather than measurement of CSF biogenic amine metabolites, better correlate with clinically important findings in schizophrenia. To test hypotheses, we used a recent technological advance in high performance liquid chromatography with electrochemical detection and combined it with multivariate statistical analyses to study biogenic amine concentrations in CSF in schizophrenia. This approach enabled the study of the interactions of several metabolites of each of the three major neurotransmitter pathways (dopaminergic, noradrenergic, and serotonergic) to test existing hypotheses regarding the neurobiochemical basis of schizophrenia. Twenty biogenic amines, their metabolites, and other compounds from 24 medication-free schizophrenic patients and 12 normal control subjects were simultaneously measured using a recently developed technique of gradient high performance liquid chromatography coupled with a 16-channel electrochemical array detector. After covariation for storage time, results of a stepwise discriminant function analysis comparing the control and patient groups identified tryptophan, tryptophol, and epinephrine as discriminating variables. Hotelling's paired T2 test from a subgroup of schizophrenic patients studied while they were and were not receiving neuroleptic treatment did not yield any significant differences between subgroups. A discussion of the findings and a comparison with previous studies of CSF biogenic amines in schizophrenia are presented.
Asunto(s)
Aminas Biogénicas/líquido cefalorraquídeo , Haloperidol/uso terapéutico , Esquizofrenia/tratamiento farmacológico , Psicología del Esquizofrénico , Adulto , Cromatografía Líquida de Alta Presión , Enfermedad Crónica , Dopamina/líquido cefalorraquídeo , Método Doble Ciego , Femenino , Humanos , Quinurenina/líquido cefalorraquídeo , Masculino , Norepinefrina/líquido cefalorraquídeo , Esquizofrenia/líquido cefalorraquídeo , Esquizofrenia/diagnóstico , Serotonina/líquido cefalorraquídeo , Triptófano/líquido cefalorraquídeo , Tirosina/líquido cefalorraquídeoRESUMEN
PURPOSE: Malignancy can be associated with high levels of catabolic products. We performed a two-part study. Part 1 measured levels of uric acid and xanthine in the aqueous humor of eyes with malignant and nonmalignant diagnoses. Part 2 measured the levels of uric acid in tears of retinoblastoma patients. If compounds in high concentrations inside the eye could be detected outside the eye, via diffusion, in high concentrations in the tears, then a tear screening test for retinoblastoma could be developed. METHODS: High-performance liquid chromatography measured levels of uric acid and xanthine in aqueous humor samples of patients with retinoblastoma, melanoma, Coats' disease, adult cataract, and congenital cataract. Tear sampling was performed on patients with retinoblastoma and on normal eyes, and samples were assayed for uric acid. RESULTS: Part 1--Uric acid was elevated in the aqueous humor of eyes with retinoblastoma, melanoma, and Coats' disease compared with eyes with cataracts. Xanthine was elevated in retinoblastoma and Coats' disease and was lower in adult and congenital cataracts and melanoma. Part 2-No significant difference was found in the concentrations of uric acid in the tears of patients with retinoblastoma and those of normal patients. CONCLUSIONS: High levels of uric acid and xanthine present in the aqueous humor of patients with malignancy are consistent with the destructive nature of these conditions. Although uric acid was not elevated in the tears of retinoblastoma patients, continued investigation into substances that might be measurably different in the tears may yield a useful screening test in the future.
Asunto(s)
Humor Acuoso/metabolismo , Melanoma/metabolismo , Neoplasias de la Retina/metabolismo , Retinoblastoma/metabolismo , Lágrimas/metabolismo , Ácido Úrico/metabolismo , Adulto , Catarata/congénito , Catarata/metabolismo , Cromatografía Líquida de Alta Presión , Humanos , Lactante , Enfermedades de la Retina/metabolismo , Xantina/metabolismoRESUMEN
The study of phytoestrogens in food sources and their metabolism, effects, and mechanism of action in animals requires very selective and often sensitive analytical techniques. We have applied coulometric array detection, which uses a series of flow-through electrochemical sensors each providing 100% electrolytic efficiency, for measurement of a variety of phytochemicals in complex matrices. Recent work has involved the resolution of coumestrol (COM), daidzein (DE), daidzin (DI), diethylstilbestrol (DES), enterodiol (ED), enterolactone (EL), equol (EQ), estradiol (E2), estriol (E3), estrone (E), genistein (GE), and quercetin (QE). Binary gradient reversed-phase (C18) chromatography was used with a sodium acetate buffer (pH 4.8)-methanol-acetonitrile solvent system. Eight coulometric sensors were set at 260, 320, 380, 440, 500, 560, 620, and 680 mV (vs Pd reference). Compounds were resolved in 30 min via both their oxidation/reduction characteristics and chromatographic behavior. Respective maximal oxidation potentials (mV) were: COM = 380; DE = 500; DI = 620; DES = 440; ED = 620; EL = 620; EQ = 560; E2 = 560; E3 = 560; E1 = 560; GE = 500; and QE = 260 with limits of detection of 5-50 pg. Uterine tissue homogenates (30 mg/ml in Tris-EDTA) and plasma from Sprague-Dawley rats sacrificed 1 hr after sc injection with either vehicle, dimethylsulfoxide, 10 microg DES, or 1.0 mg EQ were analyzed before and after enzymatic hydrolysis with beta-glucuronidase/sulfatase. Urine samples from humans receiving a Boston-area diet with or without soy protein isolate supplements were also analyzed. Ethanol extracts were evaporated and reconstituted in 20% methanol before HPLC analysis. DE, ED, EL, EQ, and GE were determined in urine with less than 5% (R.S.D.) intraassay imprecision and 85%-102% recovery. Levels (ng/ml) of GE (1.8), QE (11.2), and EQ (1.7) were found in control plasma before hydrolysis and GE (293), QE (183), and EQ (22) after hydrolysis. Higher concentrations, corresponding to sc injection, in free and total EQ were found in both tissue and plasma.
Asunto(s)
Estrógenos no Esteroides/análisis , Flavonoides , Isoflavonas , Fenoles/análisis , Polímeros/análisis , Animales , Antineoplásicos Fitogénicos/sangre , Antineoplásicos Fitogénicos/orina , Análisis Químico de la Sangre , Cromatografía Líquida de Alta Presión , Electroquímica , Estrógenos no Esteroides/sangre , Estrógenos no Esteroides/orina , Femenino , Fenoles/sangre , Fenoles/orina , Fitoestrógenos , Preparaciones de Plantas , Polifenoles , Ratas , Ratas Sprague-Dawley , Orina/químicaRESUMEN
We describe a procedure for the direct measurement of metanephrine (MN) and normetanephrine (NMN) in hydrolyzed urine, using HPLC with coulometric array detection. Acid-hydrolyzed samples were diluted and filtered before separation by isocratic reversed-phase ion-pair chromatography. Eight serial coulometric sensors, set at incrementally increasing anodic potentials, were used to screen lower-oxidizing interferences and provide stepwise oxidation of the metanephrines. Voltammetric behavior across three adjacent sensors was used to assess resolution and aid in peak identification. Values obtained in commercial controls were consistently within the specified target range. Variability, expressed as CV, was 5.45-9.22% between runs and 1.60-4.52% within-run for both compounds. The limit of detection was 2.6 micrograms/L for MN and 2.8 micrograms/L for NMN, with a linear response to 15.0 mg/L for both analytes. Results from patients' samples correlated well with those by a method involving dual ion-exchange extraction (r = 0.963, n = 82 for MN; r = 0.9768, n = 83 for NMN). This procedure provided high selectivity and objective peak purity information while greatly simplifying sample preparation.
Asunto(s)
Cromatografía Líquida de Alta Presión/métodos , Metanefrina/orina , Normetanefrina/orina , Electroquímica , Humanos , Valor Predictivo de las PruebasRESUMEN
Levels of norepinephrine, epinephrine, dopamine, and serotonin (5-HT) and their precursors [tyrosine, L-3,4-dihydroxyphenylalanine, tryptophan, and 5-hydroxytryptophan (5-HTP)] and metabolites [3,4-dihydroxyphenylacetic acid (DOPAC), 3-methoxytyramine (3-MT), homovanillic acid, 3-methoxy-4-hydroxyphenylglycol, and 5-hydroxyindoleacetic acid (5-HIAA)] were determined concurrently in samples of chick retina, pineal gland, and nine selected areas of the brain (optic lobes, thalamus, hypothalamus, optic chiasm, pons/medulla, cerebellum, neostriatum/ectostriatum, hyperstriatum, and basal forebrain) using HPLC coupled with a coulometric electrode array detection system. The norepinephrine level was highest in the pineal gland, but it was also widely distributed throughout the chick brain, with the thalamus and hypothalamus showing substantial levels. The dopamine level was highest in the basal forebrain. The epinephrine level was highest in the hypothalamus. The thalamus and hypothalamus showed the highest levels of 5-HT. Daytime levels (1100 h) of these compounds were compared with levels in chicks killed in the middle of the dark phase (2300 h). In the brain areas examined, no day/night variations in levels of norepinephrine, epinephrine, dopamine, or 5-HT were seen, although significant nocturnal changes in levels of their metabolites were observed in some areas. Pineal levels of 5-HIAA decreased significantly at night. The retina showed significant nocturnal increases in 5-HTP, 5-HT, and 5-HIAA levels. Retinal levels of 3-MT and DOPAC were significantly decreased at night.
Asunto(s)
Aminas Biogénicas/metabolismo , Encéfalo/metabolismo , Ritmo Circadiano , Glándula Pineal/metabolismo , Retina/metabolismo , Animales , Catecolaminas/metabolismo , Cromatografía Líquida de Alta Presión , Serotonina/metabolismo , Distribución TisularRESUMEN
Recent evidence suggests that there may be overactivation of the N-methyl-D-aspartate (NMDA) subtype of excitatory amino acid receptors in Huntington's disease (HD). Tryptophan metabolism by the kynurenine pathway produces both quinolinic acid, an NMDA receptor agonist, and kynurenic acid, an NMDA receptor antagonist. In the present study, multiple components of the tyrosine and tryptophan metabolic pathways were quantified in postmortem putamen of 35 control and 30 HD patients, using HPLC with 16-sensor electrochemical detection. Consistent with previous reports in HD putamen, there were significant increases in 5-hydroxyindoleacetic acid, 5-hydroxytryptophan, and serotonin concentrations. Within the kynurenine pathway, the ratio of kynurenine to kynurenic acid was significantly (p less than 0.01) increased twofold in HD patients as compared with controls, consistent with reduced formation of kynurenic acid in HD. CSF concentrations of kynurenic acid were significantly reduced in HD patients as compared with controls and patients with other neurologic diseases. Because kynurenic acid is an endogenous inhibitor of excitatory neurotransmission and can block excitotoxic degeneration in vivo, a relative deficiency of this compound could directly contribute to neuronal degeneration in HD.
Asunto(s)
Cuerpo Estriado/metabolismo , Enfermedad de Huntington/metabolismo , Ácido Quinurénico/metabolismo , Quinurenina/metabolismo , Anciano , Autopsia , Cuerpo Estriado/patología , Femenino , Humanos , Masculino , Persona de Mediana Edad , Putamen/análisis , Valores de Referencia , Triptófano/metabolismo , Tirosina/metabolismoRESUMEN
Aqueous humor from children with retinoblastoma obtained at enucleation and from eyes with adult cataracts were assayed with electrochemical liquid chromatography (Model 5500 Coulochem electrode array system) for metabolites of tyrosine, tryptophan metabolic pathways, catecholamine degradation pathways and ascorbate. More than 20 metabolites were identified in human aqueous for the first time. High levels of ascorbate were found in aqueous of eyes with adult cataracts (254, 336 ng/ml). Tyrosine metabolism in both sets of eyes was through dopamine. Vandylmandelic acid (VMA), homovanillic acid (HVA), and 3-methoxy, 4-hydroxyphenylglycol (MHPG) were all detected in retinoblastoma eyes. Although eyes with either adult cataracts or childhood retinoblastoma convert tryptophan through the serotonin pathway, retinoblastoma eyes metabolize tryptophan through the kynurenine pathway to a greater degree than adult cataract eyes.
Asunto(s)
Aminoácidos/análisis , Humor Acuoso/metabolismo , Neoplasias del Ojo/metabolismo , Retinoblastoma/metabolismo , Adulto , Ácido Ascórbico/análisis , Catarata/metabolismo , Catecolaminas/análisis , Preescolar , Cromatografía Líquida de Alta Presión , HumanosRESUMEN
Coulometric electrodes in series have been used with liquid chromatography with electrochemical detection to increase selectivity and resolution for the direct analysis of tissue neurotransmitters. Use of three coulometric sensors for electrochemical modification, selectivity, and peak identification has been expanded into "gate" cells of three or four coulometric electrodes that allow elimination of all electrochemically irreversible substances, and "array" cells of up to 15 coulometric electrodes for separation of co-eluting compounds by their current/voltage characteristics. On-column sensitivity of the sensor arrays is 0.4 to 4 pg. Gate cell selectivity favors electrochemically reversible compounds over irreversible ones, e.g., 3-methoxy-4-hydroxyphenylglycol vs ascorbate, by a factor of up to 10(4). Resolution across the multi-electrode array cells allows separation of co-eluting compounds with half-wave potentials differing by as little as 30 to 40 mV. Cells with three to 15 electrodes have been used to measure monoamines and metabolites in brain; monoamines directly in serum filtrate; and the state of oxidation of 5-hydroxytryptamine and 5-hydroxytryptophan in cerebrospinal fluid of patients with Alzheimer's disease.