RESUMEN

Opioid analgesics are clinically used to relieve severe pain in acute postoperative and cancer pain, and also in the long term in chronic pain. The analgesic action is mediated by µ-, δ-, and κ-receptors, but currently, with few exceptions for k-agonists, µ-agonists are the only ones used in therapy. Previously synthesized compounds with diazotricyclodecane cores (DTDs) have shown their effectiveness in binding opioid receptors. Fourteen novel diazatricyclodecanes belonging to the 9-propionyl-10-substituted-9,10-diazatricyclo[4.2.1.12,5]decane (compounds 20-23, 53, 57 and 59) and 2-propionyl-7-substituted-2,7-diazatricyclo[4.4.0.03,8]decane (compounds 24-27, 54, 58 and 60) series, respectively, have been synthesized and their ability to bind to the opioid µ-, δ- and κ-receptors was evaluated. Five of these derivatives, compounds 20, 21, 24, 26 and 53, showed µ-affinity in the nanomolar range with a negligible affinity towards δ- and κ-receptors and high µ-receptor selectivity. The synthesized compounds showed µ-receptor selectivity higher than those of previously reported methylarylcinnamyl analogs.

Asunto(s)

Receptores Opioides , Analgésicos , Modelos Moleculares

RESUMEN

Given that the peculiar redox behavior of ergothioneine involves a rapid regeneration process, the measurement of its precursor and redox metabolite hercynine could be particularly useful in assessing its role in oxidative stress or other biological processes. Thus, a LC-MS/MS method for the determination of hercynine concentrations in whole blood was developed. After lysis of red blood cells by cold water, samples were filtered on micro concentrators at a controlled temperature of 4 °C. The clear filtered fluid was then treated with diethylpyrocarbonate to derivatize hercynine for the analysis by LC-MS/MS. The derivatized analyte was isocratically separated as a carbethoxy derivative on a C18 column with a mobile phase of an aqueous 0.1% v/v formic acid and acetonitrile (95:5). Effluents were monitored by MRM transitions at m/z 270.28→95 and 273.21→95 for hercynine and its deuterated counterpart, respectively. No cross-talk between MRM transitions was observed and a good linearity was found within a range of 35⁻1120 nmol/L. The LOD and LOQ were, respectively, 10.30 and 31.21 nmol/L with an intraday and intermediate precision below 7%. The average hercynine concentration in whole blood from 30 healthy male volunteers (aged 77 ± 12 years) was 178.5 ± 118.1 nmol/L. Overall, the method is easy to perform, allowing a rapid and accurate assessment of whole blood concentrations of hercynine.

Asunto(s)

Antioxidantes/análisis , Betaína/análogos & derivados , Histidina/análogos & derivados , Espectrometría de Masas en Tándem/métodos , Acetonitrilos/análisis , Betaína/sangre , Cromatografía Liquida , Formiatos/análisis , Histidina/sangre , Humanos , Estrés Oxidativo/fisiología

RESUMEN

8-Chloro-1-(2',4'-dichlorophenyl)-N-piperidin-1-yl-1,4,5,6-tetrahydrobenzo[6,7]cyclohepta[1,2-c]pyrazole-3-carboxamide 9a was discovered as potent and selective CB1 antagonist by part of our group few years ago. In particular it was reported to have an affinity towards the CB1 cannabinoid receptor (CB1R), expressed as Ki, of 0.00035 nM. Nevertheless significantly divergent data were reported for the same compound from other laboratories. To unequivocally define the receptor profile of 9a, we have critically reviewed both its synthesis approach and binding data. Here we report that, in contrast to our previously reported data, 9a showed a Ki value for CB1R in the order of nanomolar rather than of fentomolar range. The new determined receptor profile of 9a was also ascertained for analogue derivatives 9b-i, as well as for 12. Moreover, the structural features of the synthesized compounds necessary for CB1R were investigated. Amongst the novel series, effects on CB1R intrinsic activity was highlighted due to the substituents at the position 3 of the pyrazole ring of the 1,4,5,6-tetrahydrobenzo[6,7]cyclohepta[1,2-c]pyrazole scaffold. Although the cannabinoid receptor profile of 9a was reviewed in this work, the relevance of this compound in CB1R antagonist based drug discovery is confirmed.

Asunto(s)

Piperidinas/síntesis química , Piperidinas/farmacología , Pirazoles/síntesis química , Pirazoles/farmacología , Receptor Cannabinoide CB1/antagonistas & inhibidores , Descubrimiento de Drogas , Unión Proteica , Relación Estructura-Actividad

RESUMEN

Here we report the investigation of diazabicycloalkane cores as potential new scaffolds for the development of novel analogues of the previously reported diazatricyclodecane selective delta (δ) opioid agonists, as conformationally constrained homologues of the reference δ agonist (+)-4-[(αR)-α((2S,5R)-4-allyl-2,5-dimethyl-1-piperazinyl)-3-methoxybenzyl]-N,N-diethylbenzamide (SNC80). In particular, we have simplified the diazatricyclodecane motif of δ opioid agonist prototype 1a with bridged bicyclic cores. 3,6-diazabicyclo[3.1.1]heptane, 3,8-diazabicyclo[3.2.1]octane, 3,9-diazabicyclo[3.3.1]nonane, 3,9-diazabicyclo[4.2.1]nonane, and 3,10-diazabicyclo[4.3.1]decane were adopted as core motifs of the novel derivatives. The compounds were synthesized and biologically assayed as racemic (3-5) or diastereoisomeric (6,7) mixtures. All the novel compounds 3-7 showed δ agonism behaviour and remarkable affinity to δ receptors. Amongst the novel derivatives, 3,8-diazabicyclo[3.2.1]octane based compound 4 evidenced improved δ affinity and selectivity relative to SNC80.

Asunto(s)

Alcanos/química , Receptores Opioides delta/agonistas , Receptores Opioides mu/agonistas , Estructura Molecular , Relación Estructura-Actividad

RESUMEN

Novel pyrazolo[5,1-f][1,6]naphthyridines, pyrazolo[5,1-a][2,6]naphthyridines, pyrazolo[5,1-a][2,7]naphthyridines and pyrazolo[5,1-a]isoquinolines phenylimidazole/benzimidazole ethylene-linked were designed and synthesized for PDE10A interaction. An AgOTf and proline-cocatalyzed multicomponent methodology based on use of o-alkynylaldehydes, tosylhydrazide and ketones was developed and proved to be a convenient route for assembly of most of the novel tricyclic pyrazoles synthesized. Pyrazolo[5,1-f][1,6]naphthyridine 43 and 59, pyrazolo[5,1-a][2,6]naphthyridine 66, and pyrazolo[5,1-a][2,7]naphthyridine 42 showed the highest affinity for PDE10A enzyme (IC50 = 40, 42, 40, 55 nM, respectively).

Asunto(s)

Inhibidores de Fosfodiesterasa/síntesis química , Inhibidores de Fosfodiesterasa/farmacología , Hidrolasas Diéster Fosfóricas/metabolismo , Pirazoles/farmacología , Animales , Relación Dosis-Respuesta a Droga , Humanos , Microsomas Hepáticos/efectos de los fármacos , Microsomas Hepáticos/enzimología , Microsomas Hepáticos/metabolismo , Modelos Moleculares , Estructura Molecular , Inhibidores de Fosfodiesterasa/química , Pirazoles/síntesis química , Pirazoles/química , Ratas , Relación Estructura-Actividad

RESUMEN

Considering the interesting pharmacological profile of the delta (δ) selective opioid agonist compound SNC-80, conformationally constrained analogs containing two diazatricyclodecane ring systems in place of dimethylpiperazine core motif were synthesized. The compounds showed subnanomolar or low nanomolar δ opioid receptor binding affinity. Depending upon the substituents on the diazatricyclodecane ring, these compounds displayed varying selectivity for δ opioid receptor over µ and κ receptors. Amongst the novel compounds, 1Aa showed the more interesting biological profile, with higher δ affinity and selectivity compared to SNC-80. The δ receptor agonist profile and antinociceptive activity of 1Aa were confirmed using ex-vivo (isolated mouse vas deferens) and in vivo (tail flick) assays.

Asunto(s)

Analgésicos/farmacología , Dolor/tratamiento farmacológico , Compuestos Policíclicos/farmacología , Receptores Opioides delta/agonistas , Analgésicos/administración & dosificación , Analgésicos/síntesis química , Animales , Relación Dosis-Respuesta a Droga , Ligandos , Ratones , Estructura Molecular , Dimensión del Dolor , Compuestos Policíclicos/administración & dosificación , Compuestos Policíclicos/síntesis química , Relación Estructura-Actividad

RESUMEN

Two sensitive and reproducible capillary electrophoresis and high-performance liquid chromatography-fluorescence procedures were established for quantitative determination of L-egothioneine in plasma. After derivatization of L-ergothioneine with 5-iodoacetamidofluorescein, the separation was carried out by HPLC on an ODS-2 C-18 sperisorb column by using a linear gradient elution and by HPCE on an uncoated fused silica capillary, 50 µm id, and 60 cm length. The methods were validated and found to be linear in the range of 0.3 to 10 µmol/l. The limit of quantification was 0.27 µmol/l for HPCE and 0.15 µmol/l for HPLC. The variations for intra- and inter-assay precision were around 6 RSD%, and the mean recovery accuracy close to 100% (96.11%).

Asunto(s)

Ergotioneína/sangre , Cromatografía Líquida de Alta Presión , Electroforesis Capilar , Ergotioneína/química , Fluoresceínas/química , Humanos , Reproducibilidad de los Resultados , Sensibilidad y Especificidad , Espectrometría de Fluorescencia

RESUMEN

For the past few decades membrane zinc metallopeptidases have been identified as important therapeutic targets in the control of pain. In particular, neutral endopeptidase (NEP) has been shown to play critical roles in the metabolism of the endogenous peptides Met- and Leu-enkephalins. In this study, we have evaluated the activity of a new fluorinated peptidase inhibitor NESS002ie in both in vitro and in vivo assays. NESS002ie has been compared to the peptidomimetic compound thiorphan and the previously reported NEP selective thiol inhibitor C20. The metallopeptidases inhibitory activity of NESS002ie was tested in vitro using a highly, sensitive, continuous, fluorometric, enzyme assay. Also, the analgesic propriety of NESS002ie, thiorphan and C20 have been evaluated in vivo, by intraplantar, intravenous and intrathecal administration, through nociception assays based on formalin test in mice. Metallopeptidases assays have shown an inhibitory potency of NESS002ie in the nanomolar range for NEP and angiotensin-converting enzyme (ACE). The new fluorinated inhibitor showed higher analgesic activity and bioavailability compared to thiorphan and C20 when administered by both intravenous and intrathecal injections. More significantly, intrathecal injection of NESS002ie reduced both the first and the second phases of the formalin biphasic pain response. In addition, naltrindole and naloxone reversed the analgesic effect of NESS002ie with a diverse profile. This study shows an improvement in relief of inflammation and pain, in vivo, using NESS002ie compared to reference compounds thiorphan and C20. This significant effect could be due to the replacement of isobutyl chain of the thiol C20 with the trifluoromethyl group.

Asunto(s)

Analgésicos/farmacología , Modelos Animales de Enfermedad , Dolor Intratable/tratamiento farmacológico , Inhibidores de Proteasas/farmacología , Compuestos de Sulfhidrilo/farmacología , Tirosina/análogos & derivados , Analgésicos/uso terapéutico , Animales , Formaldehído/administración & dosificación , Masculino , Ratones , Naloxona/administración & dosificación , Inhibidores de Proteasas/uso terapéutico , Compuestos de Sulfhidrilo/uso terapéutico , Tirosina/farmacología , Tirosina/uso terapéutico

RESUMEN

We have recently synthesized a new series of 4,5-dihydrobenzo-oxa-cycloheptapyrazole derivatives with the aim to discover novel CB1 antagonist agents characterized by anti-obesity activity comparable to that of SR141716A but with reduced adverse effects such as anxiety and depression. Within the novel class, the CB1 antagonist 8-chloro-1-(2,4-dichlorophenyl)-N-piperidin-1-yl-4,5-dihydrobenzo-1H-6-oxa-cyclohepta(1,2-c)pyrazole-3-carboxamide (NESS06SM) has been selected as lead compound. We found that NESS06SM is a CB1 neutral antagonist, characterized by poor blood-brain barrier permeability. Moreover, NESS06SM chronic treatment determined both anti-obesity effect and cardiovascular risk factor improvement in C57BL/6N Diet Induced Obesity (DIO) mice fed with fat diet (FD mice). In fact, the mRNA gene expression in Central Nervous System (CNS) and peripheral tissues by real time PCR, showed a significant increase of orexigenic peptides and a decrease of anorexigenic peptides elicited by NESS06SM treatment, compared to control mice fed with the same diet. Moreover, in contrast to SR141716A treatment, the chronic administration of NESS06SM did not change mRNA expression of both monoaminergic transporters and neurotrophins highly related with anxiety and mood disorders. Our results suggest that NESS06SM reduces body weight and it can restore the disrupted expression profile of genes linked to the hunger-satiety circuit without altering monoaminergic transmission probably avoiding SR141716A side effects. Therefore the novel CB1 neutral antagonist could represent a useful candidate agent for the treatment of obesity and its metabolic complications.

Asunto(s)

Fármacos Antiobesidad/uso terapéutico , Benzoxepinas/uso terapéutico , Antagonistas de Receptores de Cannabinoides/uso terapéutico , Obesidad/tratamiento farmacológico , Pirazoles/uso terapéutico , Animales , Fármacos Antiobesidad/farmacología , Benzoxepinas/farmacología , Peso Corporal/efectos de los fármacos , Antagonistas de Receptores de Cannabinoides/farmacología , Dieta Alta en Grasa , Ácido Graso Sintasas/genética , Glucoquinasa/genética , Hígado/efectos de los fármacos , Hígado/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Obesidad/metabolismo , Piperidinas/farmacología , Piperidinas/uso terapéutico , Pirazoles/farmacología , Piruvato Quinasa/genética , ARN Mensajero/metabolismo , Receptor Cannabinoide CB1/genética , Receptor Cannabinoide CB1/metabolismo , Rimonabant , Simportadores/genética

RESUMEN

A new hydrophilic interaction ultra-performance LC method was established for the whole blood measurement of L-ergothioneine. Chromatographic separation was achieved in a fairly short time, less than 4 min, on a 100 × 2.1 mm Acquity UPLC BEH HILIC 1.7 µm column with a mobile phase consisting of a mixture of 100 mmol/L ammonium acetate/ACN/water (5:85:10, v/v/v) that flowed isocratically at 0.250 mL/min. The LOD and the limit of quantification were 3.85 and 11.67 µmol/L, respectively. The method exhibited linearity in a concentration range of 15.63-1000 µmol/L (R(2) > 0.999). Mean recovery was 96.34% whereas intraassay and interassay precision were 1.52 and 1.82% RSD, respectively. On the whole, the developed method is simple, fast, precise, accurate, and sensitive and may be useful for routine analyses.

Asunto(s)

Cromatografía Líquida de Alta Presión/métodos , Ergotioneína/sangre , Anciano , Anciano de 80 o más Años , Cromatografía Líquida de Alta Presión/instrumentación , Femenino , Humanos , Interacciones Hidrofóbicas e Hidrofílicas , Masculino , Espectrofotometría Ultravioleta

RESUMEN

In spite of rimonabant's withdrawal from the European market due to its adverse effects, interest in the development of drugs based on CB1 antagonists is revamping on the basis of the peculiar properties of this class of compounds. In particular, new strategies have been proposed for the treatment of obesity and/or related risk factors through CB1 antagonists, i.e. by the development of selectively peripherally acting agents or by the identification of neutral CB1 antagonists. New compounds based on the lead CB1 antagonist/inverse agonist rimonabant have been synthesized with focus on obtaining neutral CB1 antagonists. Amongst the new derivatives described in this paper, the mixture of the two enantiomers (±)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-3-(2-cyclohexyl-1-hydroxyethyl)-4-methyl-1H-pyrazole ((±)-5), and compound 5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-3-[(Z)-2-cyclohexyl-1-fluorovinyl]-4-methyl-1H-pyrazole ((Z)-6), showed interesting pharmacological profiles. According to the preliminary pharmacological evaluation, these novel pyrazole derivatives showed in fact both neutral CB1 antagonism behaviour and significant in vivo activity towards food intake.

Asunto(s)

Ingestión de Alimentos/efectos de los fármacos , Pirazoles/farmacología , Receptor Cannabinoide CB1/antagonistas & inhibidores , Animales , Relación Dosis-Respuesta a Droga , Masculino , Ratones , Estructura Molecular , Pirazoles/síntesis química , Pirazoles/química , Relación Estructura-Actividad

RESUMEN

The present work aims to study the effects induced by a chronic treatment with a novel CB1 antagonist (NESS038C6) in C57BL/6N diet-induced obesity (DIO) mice. Mice treated with NESS038C6 and fed with a fat diet (NESS038C6 FD) were compared with the following three reference experimental groups: DIO mice fed with the same fat diet used for NESS038C6 and treated with vehicle or the reference CB1 antagonist/inverse agonist rimonabant, "VH FD" and "SR141716 FD", respectively; DIO mice treated with vehicle and switched to a normal diet (VH ND). NESS038C6 chronic treatment (30 mg/kg/day for 31 days) determined a significant reduction in DIO mice weight relative to that of VH FD. The entity of the effect was comparable to that detected in both SR141716 FD and VH ND groups. Moreover, if compared to VH FD, NESS038C6 FD evidenced: (i) improvement of cardiovascular risk factors; (ii) significant decrease in adipose tissue leptin expression; (iii) increase in mRNA expression of hypothalamic orexigenic peptides and a decrease of anorexigenic peptides; (iv) expression increase of metabolic enzymes and peroxisome proliferator-activated receptor-α in the liver; (v) normalization of monoaminergic transporters and neurotrophic expression in mesolimbic area. However, in contrast to the case of rimonabant, the novel CB1 antagonist improved the disrupted expression profile of genes linked to the hunger-satiety circuit, without altering monoaminergic transmission. In conclusion, the novel CB1 antagonist compound NESS038C6 may represent a useful candidate agent for the treatment of obesity and its metabolic complications, without or with reduced side effects relative to those instead observed with rimonabant.

Asunto(s)

Fármacos Antiobesidad/uso terapéutico , Antagonistas de Receptores de Cannabinoides/uso terapéutico , Obesidad/tratamiento farmacológico , Receptor Cannabinoide CB1/antagonistas & inhibidores , Receptor Cannabinoide CB1/metabolismo , Análisis de Varianza , Animales , Glucemia , Índice de Masa Corporal , Peso Corporal/efectos de los fármacos , Factor Neurotrófico Derivado del Encéfalo/genética , Factor Neurotrófico Derivado del Encéfalo/metabolismo , Antagonistas de Receptores de Cannabinoides/química , Colesterol/sangre , Grasas de la Dieta/efectos adversos , Modelos Animales de Enfermedad , Proteínas de Transporte de Dopamina a través de la Membrana Plasmática/genética , Proteínas de Transporte de Dopamina a través de la Membrana Plasmática/metabolismo , Relación Dosis-Respuesta a Droga , Regulación de la Expresión Génica/efectos de los fármacos , Hipotálamo/efectos de los fármacos , Hipotálamo/metabolismo , Indazoles/farmacología , Indazoles/uso terapéutico , Grasa Intraabdominal/patología , Hígado/efectos de los fármacos , Hígado/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Factor de Crecimiento Nervioso/genética , Factor de Crecimiento Nervioso/metabolismo , Obesidad/sangre , Obesidad/inducido químicamente , Piperidinas/química , Piperidinas/uso terapéutico , Pirazoles/química , Pirazoles/uso terapéutico , Receptor Cannabinoide CB1/genética , Rimonabant , Proteínas de Transporte de Serotonina en la Membrana Plasmática/genética , Proteínas de Transporte de Serotonina en la Membrana Plasmática/metabolismo , Tiofenos/farmacología , Tiofenos/uso terapéutico , Transaminasas/sangre , Triglicéridos/sangre

RESUMEN

AIM: Different studies, carried out by us and others, have investigated the impact of carbon nanotubes (CNTs) in vitro and in animal models. To date, only a few studies have been performed on human cells ex vivo. There is also a lack of comparison between CNTs with varied functionalization and structural properties and their impact on different cell types. MATERIALS & METHODS: The present ex vivo human study focuses on the impact of a series of functionalized multiwalled CNTs on human T and B lymphocytes, natural killer (NK) cells and monocytes. RESULTS: Smaller diameter nanotubes are internalized more efficiently. Viability assays displayed the absence of cytotoxicity of all multiwalled CNTs used. Activation assay demonstrated a strong effect on monocytes and NK cells. CONCLUSION: Our results, on human cells ex vivo, confirmed previous studies demonstrating appropriately functionalized CNTs are nontoxic. The effects on cell functionality were significant for the monocytes and NK cells. These findings encourage the possible use of CNTs for biomedical applications either as carriers of therapeutic molecules or as immune modulator systems.

Asunto(s)

Citocinas/inmunología , Inmunidad Innata/efectos de los fármacos , Inmunidad Innata/inmunología , Nanotubos de Carbono/toxicidad , Linfocitos T/efectos de los fármacos , Linfocitos T/inmunología , Células Cultivadas , Humanos , Ensayo de Materiales

RESUMEN

An unusual reaction involving ninhydrin and aminothiols was exploited to set an indirect method for the chiral recognition of stereoisomers of penicillamine. Separation of diastereoisomers was achieved on a C18 column in isocratic mode by using a mixture of propionic acid (pH 3.0)/acetonitrile/water (10:10:80, v/v/v) as a mobile phase. Diastereoisomers were detected by a fluorescence detector in fairly short times (about 7 min) and with a good resolution. The lowest detectable amount of toxic isomer of penicillamine (l-enantiomer) in samples of the d-enantiomer, was around 0.01%. The method was also suitable for the indirect chiral recognition of other aminothiols such as cysteine and cysteinylglycine.

Asunto(s)

Cromatografía Líquida de Alta Presión/métodos , Cromatografía de Fase Inversa/métodos , Ninhidrina/química , Penicilamina/química , Penicilamina/aislamiento & purificación , Cisteína/química , Estereoisomerismo

Asunto(s)

Apolipoproteínas B/metabolismo , Cisteína/metabolismo , Degeneración Hepatolenticular/metabolismo , Homocisteína/metabolismo , Penicilamina/metabolismo , Degeneración Hepatolenticular/tratamiento farmacológico , Humanos , Penicilamina/sangre , Penicilamina/uso terapéutico , Compuestos de Sulfhidrilo/sangre

RESUMEN

A hydrophilic interaction chromatography-based method, in combination with 1.7 microm ethylene bridged hybrid particle packed column (100 mm x 2.1 mm I.D.) and ultraperformance liquid chromatography, has been developed to measure cytosine (C) and methylcytosine (mC) in order to evaluate the extent of DNA methylation. Separation of cytosine and methylcytosine was achieved with good resolution and in fairly short times (5.5 min) by using isocratic elution with a mixture of 97:3 (v/v) acetonitrile/10 mM ammonium acetate as a mobile phase. The determination coefficients of C and mC were high (R(2) > 0.999) within the range tested. The %RSD for intraday and interday were respectively 2.2% and 2.5% for C and 3.5% and 3.8% for mC. The limit of detection was 0.52 microM (0.52 fmol on-column) both for C and mC while the limit of quantification was 1.72 microM (1.72 fmol on-column) both for C and mC. The smallest amount of purified DNA that yielded a measurable level of C and mC was 10 microg. On the whole, this method is simple, rapid, sensitive, and precise.

Asunto(s)

Cromatografía Liquida/métodos , Metilación de ADN , Genoma Humano , Espectrofotometría Ultravioleta/métodos , Animales , ADN/análisis , Humanos , Agua/química

RESUMEN

A rapid and simple short-end injection capillary zone electrophoresis method was developed for the quantification of plasma uric acid. The separation was performed in an uncoated fused-silica capillary (50 microm ID, 60 cm total length, 10.2 cm effective length) by using as a background electrolyte a 75 mmol/L glycylglycine solution titrated with NaOH 5 mol/L to pH 9.0, a voltage of 28 kV, a cartridge temperature of 15 degrees C, and direct UV detection at 292 nm. Under optimized conditions, uric acid was determinate in little more than 1 min (1.076 minutes). In order to verify the accuracy of the analysis, urate levels were measured in 543 apparently healthy volunteers by the new assay and our previous method, and the obtained data were compared by Passing-Bablock regression, Bland-Altman test, and a new regression-based approach, which showed a good agreement between two methods.

Asunto(s)

Bioensayo/métodos , Electroforesis Capilar/métodos , Humanos , Ácido Úrico/sangre

RESUMEN

A new HPLC method by fluorescence or UV/vis absorbance detection has been developed for the separation and quantification of penicillamine stereoisomers after their spirocyclization with ninhydrin. The separation was performed on an achiral C18 column by isocratic elution with a copper(II)-l-proline complex as a chiral selector in the mobile phase. The method was able to detect traces of l-penicillamine in samples of d-penicillamine below 0.1% in fairly short times (about 16 min) with a good resolution (R(s)=1.31). On the whole, the method was found to be stable and useful in the quality control of the bulk material and formulations.

Asunto(s)

Cromatografía Líquida de Alta Presión/métodos , Penicilamina/aislamiento & purificación , Cobre/química , Ninhidrina/química , Penicilamina/análisis , Penicilamina/química , Prolina/química , Compuestos de Espiro/síntesis química , Estereoisomerismo

RESUMEN

BACKGROUND: A scatter plot combined with correlation and regression analysis to assess the agreement between analytical methods that measure the same quantity has a long tradition in clinical chemistry. However, only when linearity is absolute this approach may be useful to estimate whether the methods under comparison produce the same errors in measurement. The aim of this study is to provide a new original regression-based tool which does not suffer from the problems of the classical regression/correlation method and that is also easy to use and to interpret. METHODS: To evaluate the performance of the new method, the results of some analytical determinations were compared by the new procedure, the Bland-Altman test, Passing-Bablok regression and Mountain plot. RESULTS: All the tests used to compare the data allowed superimposable interpretations, both when methods under comparison were in agreement and in disagreement. CONCLUSIONS: The understanding of the new test is truly intuitive and it is reduced to the appraisal of how much the equation of the best-fit line (Y=X) is able to predict the value of the Y. The goodness of this prediction can be evaluated by the coefficient of determination, whose value is closely connected to the differences between the methods under comparison. The result of our simulation is that quite high agreement between the methods occurs for determination coefficient equal or greater than 0.98.

Asunto(s)

Arginina/análogos & derivados , Pruebas de Química Clínica , Péptido Natriurético Encefálico/análisis , Arginina/análisis , Cromatografía Líquida de Alta Presión , Pruebas de Química Clínica/normas , Pruebas de Química Clínica/estadística & datos numéricos , Electroforesis Capilar , Ensayo de Inmunoadsorción Enzimática , Estudios de Evaluación como Asunto , Humanos , Ensayo Inmunorradiométrico , Análisis de Regresión

RESUMEN

We report a new selective and direct pre-column ninhydrin-based derivatization reaction for determination of plasma ADMA levels. This original derivatization procedure matched to a validated and rapid RP-HPLC method can be a useful alternative to other assays in which time consuming and expensive extraction and/or purification steps are required.

Asunto(s)

Arginina/análogos & derivados , Cromatografía Líquida de Alta Presión/métodos , Ninhidrina/química , Adulto , Anciano , Anciano de 80 o más Años , Arginina/sangre , Arginina/química , Femenino , Fluorescencia , Humanos , Masculino , Persona de Mediana Edad , Sensibilidad y Especificidad , Espectrofotometría Ultravioleta/métodos , Factores de Tiempo