Search

A method was established for the simultaneous determination of 3-methoxytyramine and adrenaline in dairy products by high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) compled with isotope dilution. Sample was extracted by perchloric acid solution, cleaned by weak cationic solid phase extraction column. The analyte was separated by ACQUITY UPLC BEH C18 (100 mm×2.1 mm, 1.7 μm) column and agradient elution of mobile phase with 0.1% formic acid aqueous solution-methanol. The detection was achieved by HPLC-MS/MS with electrospray ion source and positive ion scanning mode as well as multiple reaction monitoring (MRM) mode. The quantification was accomplished by the internal standard method. It was demonstrated that the linear relationships of 3-methoxytyramine and adrenaline were good in the range of 0.5~12.0 ng/mL and 50~1200 ng/mL (r>0.99), respectively. The detection limits and the limit of quantification of 3-methoxytyramine were 0.2 and 0.5 μg/kg, respectively. At the concentrations of 0.5, 1.0 and 3.0 μg/kg, the recoveries of the 3-methoxytyramine ranged from 76.97% to 119.20%, with the corresponding precision (n=6) of 1.53%~9.61%. While the recoveries of adrenaline at 50, 100 and 300 μg/kg were 90.65%~108.55% and the relative standard deviations were in the range of 1.29%~3.95%. The method would be accurate in results and good in repeatability, and it can be an important measure to monitor 3-methoxytyramine and adrenaline in dairy products.

Background: Bilateral carotid body tumors with a concomitant skull-base paraganglioma are extremely rare, of which only one case has been reported in the literature to date. Case presentation: We present the case of a 35-year-old male with 1 year of hypertension and high levels of dopamine and 3-methoxytyramine. Magnetic resonance imaging (MRI) scans demonstrated three separate masses at the left middle cranial fossa floor and bilateral carotid bifurcation. Genetic testing showed succinate dehydrogenase complex subunit D mutation. The patient underwent the resection of the left skull base mass. Histopathology and immunohistochemistry confirmed the presence of a skull-base paraganglioma. Conclusions: Succinate dehydrogenase complex subunit D mutation-associated bilateral carotid body tumors with a concomitant skull-base paraganglioma accompanied by abnormal dopamine and hypertension are extremely rare, which not only provides ideas for considering the association of gene mutations, biochemical abnormalities and clinical symptoms but also provides an expanded diagnostic spectrum for paraganglioma in atypical locations. [ABSTRACT FROM AUTHOR]

A reliable and robust liquid chromatography–tandem mass spectrometry (LC–MS/MS) method was developed in this study for clinical testing of 24-h urinary fractionated metanephrines (MNs) and 3-methoxytyramine. At first, the acid hydrolysis condition was well optimized. Following solid-phase extraction (SPE) was performed to purify urine specimens, and the obtained elutes were directly used for LC–MS/MS analysis. A total run time of 3 min for each sample made it possible to analyze more than 300 samples per day. This method was validated with good linearity (linear correlation coefficient ≥ 0.9969), favorable precision (the coefficients of variation ≤ 10.2%), satisfactory recovery (85.0%–108%) and negligible matrix effect (92.0%–109%). The reference intervals of these analytes in Chinese subjects were established for clinical application in diagnosis of pheochromocytoma. This reliable and well-validated measurement method for 24-h urinary fractionated MNs and 3-methoxytyramine using LC–MS/MS could be successful used in clinical testing for diagnosis of pheochromocytoma. [ABSTRACT FROM AUTHOR]

A novel dual-template magnetic molecularly imprinted polymer (MMIP) was synthesized to extract normetanephrine (NMN), metanephrine (MN) and 3-methoxytyramine (3-MT) from spot urine samples. As the adsorbent of dispersive solid-phase extraction (d-SPE), the MMIP was prepared using dopamine and MN as dual templates, methacrylic acid as the functional monomer, ethylene glycol dimethacrylate as the crosslinking reagent and magnetic nanoparticles as the magnetic core. NMN, MN, 3-MT and creatinine (Cr) in spot urine samples were selectively enriched by d-SPE and detected by HPLC-fluorescence detection/ultraviolet detection. The peak area (A) ratios of NMN, MN and 3-MT to Cr were used for the diagnosis of pheochromocytomas and paragangliomas (PPGLs). The results showed that the adsorption efficiencies of MMIP for target analytes were all higher than 89.0%, and the coefficient variation precisions of intra-assay and inter-assay for the analytes were within 4.9% and 6.3%, respectively. The recoveries of the analytes were from 93.2% to 112.8%. The MMIP was still functional within 14 days and could be reused at least seven times. The d-SPE and recommended solid-phase extraction (SPE) were both used to pretreat spot urine samples from 18 PPGLs patients and 22 healthy controls. The correlation coefficients of ANMN/ACr and AMN/ACr between d-SPE and SPE were both higher than 0.95. In addition, the areas under the receiver operator curves for spot urine ANMN/ACr, AMN/ACr and plasma free NMN and MN were 0.975, 0.773 and 0.990, 0.821, respectively, indicating the two methods had the similar performances. The d-SPE method took only 20 min, which was effective in clinical application. [ABSTRACT FROM AUTHOR]

Equine urine analysis has evolved over time to detect thousands of urinary compounds for doping control in the horse racing industry. The longitudinal assessment of 3‐methoxytyramine to tyramine ratio (3‐MT/T) values in equine urine by GC–MS profiling was investigated to support the Racing NSW Equine Biological Passport (EBP) for detection of dopaminergic manipulation in racehorses. This involved comparison of routine urine samples to administration studies of Sinemet, a common Parkinson's disease medication containing levodopa. Using an endogenous reference compound (ERC) in a urinary ratio enabled greater confidence to provide intelligence of pharmaceutical manipulation as distinct from physiological variation. Population reference limits (PRLs) of 776 ng/ml for urinary 3‐MT and 5.3 for 3‐MT/T, together with the use of individual reference limits (IRLs), are proposed. [ABSTRACT FROM AUTHOR]

The abuse of performance‐enhancing catecholamine‐based stimulants, such as levodopa, is controlled in horse racing through the application of a regulatory threshold for the common major metabolite. However, catechol‐O‐methyltransferase (COMT) enzyme inhibitors can be used to restrict the catalysis of the stimulant, and so the concurrent administration of both substances would be a viable strategy to enhance racing performance while removing the risk of exceeding the threshold. A 200 mg dose of nitecapone, a COMT inhibitor, was administered to a Thoroughbred horse, and we have analysed the blood (≤24 h) and urine (≤48 h) samples that were collected. The extracts, analysed by UHPLC coupled to a high‐resolution accurate mass spectrometer, were consistent with the presence of nitecapone glucuronide in all the samples collected. An in‐depth examination of the samples was then carried out using targeted accurate mass extracted ion chromatograms to identify whether the metabolites that have been found in other species were also present in the extracts. Once these were tentatively identified, MS/MS experiments were conducted on some of the metabolites (M1–M5), as well as decomposition products (DP1 and DP2), to verify that spectrum included MS fragments were consistent with their proposed structures. The accumulated data provided evidence that is consistent with this drug having been converted into many metabolites and a few decomposition products. An unexpected finding was that O‐methylation was a very minor pathway until after the reduction of the 2,4‐pentanedione side chain had occurred. [ABSTRACT FROM AUTHOR]

Objective: Measurements of plasma free metanephrines (MNs), including MN and normetanephrine, provide high sensitivity and specificity for the diagnosis of pheochromocytoma and paraganglioma (PPGL). 3‐Methoxytyramine (3‐MT) and chromogranin A (CgA) may allow the detection of dopamine‐producing or biochemically silent PPGL. The aim of this study was to evaluate whether measurements of plasma 3‐MT or CgA as a supplement of plasma MNs offer a better diagnostic strategy for initial testing of PPGL. Patients and Design: We enroled 125 patients who underwent surgery from 2015 to 2016 for our study and identified 33 patients with PPGL and 92 patients with non‐PPGL masses. Measurement: The levels of plasma free MNs and 3‐MT were measured for all 125 patients using liquid chromatography–tandem mass spectrometry. Plasma CgA concentrations were determined using a radioimmunoassay. To evaluate the diagnostic performance of plasma free MNs, 3‐MT and CgA, sensitivity and specificity were determined, and receiver operating characteristic curves were constructed. Results: We found that combining 3‐MT and MNs increased the diagnostic sensitivity from 93.9% (95% confidence interval [CI]: 78.4%–98.9%) to 97.0% (95% CI: 82.5%–99.8%). In contrast, addition of plasma CgA test reduced the diagnostic specificity significantly from 91.3% (95% CI: 83.1%–95.9%) to 75.0% (95% CI: 64.7%–83.2%). Conclusion: Here, we demonstrated that 3‐MT represents a valuable supplementary test to plasma MNs, which can further enhance the sensitivity of the assay, while plasma CgA added no additional diagnostic value to MNs due to the lowered diagnostic specificity. [ABSTRACT FROM AUTHOR]

Dementia is exacerbated by loss of appetite and amotivation, and recent studies have indicated that ninjin'yoeito improves anorexia and amotivation. Previous studies suggest that ninjin'yoeito inhibits dopamine-metabolizing enzymes and enhances dopamine signaling. However, whether ninjin'yoeito increases dopamine content in living cells remains unclear. Here, PC12 cells were used to examine whether ninjin'yoeito affects the dopamine metabolic pathway. Dopamine content significantly increased 3 h after treatment ninjin'yoeito extract. Concomitantly, the levels of 3-methoxytyramine and 3,4-dihydroxyphenylacetic acid were significantly reduced. The effects of components of ninjin'yoeito on the dopamine metabolic pathway were also assessed. Treatment with onjisaponin B, nobiletin, and schisandrin, and the ingredients of Polygalae Radix, Citri Unshiu Pericarpium, and Schisandrae Fructus increased dopamine content and decreased its metabolite content in the culture media. Our findings suggest that ninjin'yoeito improves anorexia and amotivation by inhibiting metabolic enzyme and increasing the dopamine content in cells. Three components in NYT reduce dopamine metabolites and increase dopamine levels. [ABSTRACT FROM AUTHOR]

Accurate diagnosis of pheochromocytoma and paraganglioma (PPGLs) is highly dependent on the detection of metanephrines and catecholamines. However, the systematic investigation on influencing factors including specimen (plasma or whole blood), anticoagulant, storage conditions, and interference factors need further confirmation. Blood with heparin-lithium or EDTA-K2 were collected, stability of epinephrine (EPI), norepinephrine (NE), dopamine (DA), metanephrine (MN), normetanephrine (NMN), 3-methoxytyramine (3-MT) in whole blood and plasma at room temperature and 4 °C for different storage times, stability of plasma MN, NMN and 3-MT at −20 °C and −80 °C were investigated. Plasma with hemoglobin (1 g/L, 2 g/L, 3 g/L, 4 g/L, 6 g/L), TG (<5 mmol/L, 5–8 mmol/L, >8 mmol/L) were prepared. EPI, NE, DA were prone to degrade at room temperature, samples should be centrifuged at 4 °C. EPI and NE were stable in whole blood at 4 °C for 4 h and in plasma for 2 h. For MN, NMN, 3-MT, plasma can be stable at room temperature and 4 °C for at least 6 h, which is better than whole blood; there was no significant difference when stored at −20 °C and −80 °C for 7 days. Heparin-lithium had a slight advantage over EDTA-K2. EPI, NE, DA should not be performed when Hb > 1 g/L or TG > 5 mmol/L. MN, NMN, 3-MT should not be performed when Hb > 2 g/L, whereas TG had no interference. According to the actual clinical application scenario, this study provided a reliable basis for the accurate diagnosis of PPGLs. [ABSTRACT FROM AUTHOR]

A novel dual-template magnetic molecularly imprinted polymer (MMIP) was synthesized to extract normetanephrine (NMN), metanephrine (MN) and 3-methoxytyramine (3-MT) from spot urine samples. As the adsorbent of dispersive solid-phase extraction (d-SPE), the MMIP was prepared using dopamine and MN as dual templates, methacrylic acid as the functional monomer, ethylene glycol dimethacrylate as the crosslinking reagent and magnetic nanoparticles as the magnetic core. NMN, MN, 3-MT and creatinine (Cr) in spot urine samples were selectively enriched by d-SPE and detected by HPLC-fluorescence detection/ultraviolet detection. The peak area (A) ratios of NMN, MN and 3-MT to Cr were used for the diagnosis of pheochromocytomas and paragangliomas (PPGLs). The results showed that the adsorption efficiencies of MMIP for target analytes were all higher than 89.0%, and the coefficient variation precisions of intra-assay and inter-assay for the analytes were within 4.9% and 6.3%, respectively. The recoveries of the analytes were from 93.2% to 112.8%. The MMIP was still functional within 14 days and could be reused at least seven times. The d-SPE and recommended solid-phase extraction (SPE) were both used to pretreat spot urine samples from 18 PPGLs patients and 22 healthy controls. The correlation coefficients of ANMN/ACr and AMN/ACr between d-SPE and SPE were both higher than 0.95. In addition, the areas under the receiver operator curves for spot urine ANMN/ACr, AMN/ACr and plasma free NMN and MN were 0.975, 0.773 and 0.990, 0.821, respectively, indicating the two methods had the similar performances. The d-SPE method took only 20 min, which was effective in clinical application.

Availability of appropriately established reference intervals for biochemical tests can be troublesome in pediatrics. Here we establish age-specific continuous reference intervals for catecholamine O -methylated metabolites in children evaluated for catecholamine producing tumors, particularly younger children with suspected neuroblastoma. Plasma concentrations of 3-methoxytyramine, normetanephrine, metanephrine, and 3- O -methyldopa were analyzed by liquid chromatography tandem mass spectrometry in 533 children aged 2 days to 18 years. Concentrations of plasma free normetanephrine, 3-methoxytyramine and 3- O -methyldopa were higher in neonates up until six months of age, but thereafter declined steeply to levels after one year that were <38% those of neonatal concentrations and to further lower concentrations in teenagers that were <23% those in neonates. In contrast, concentrations of plasma free metanephrine showed a reciprocal pattern with 50% lower concentrations in infants below one year compared to later in childhood. The dynamic reciprocal changes in plasma concentrations of normetanephrine, 3-methoxytyramine and 3- O -methyldopa compared to metanephrine during early childhood suggest underlying developmental changes in extra-adrenal and adrenal chromaffin tissue that must be considered for pediatric reference intervals, particularly in infants. With such reference intervals at hand, biochemical testing for catecholamine producing tumors in young children is substantially improved. • LC-MS/MS based measurements of plasma catecholamine metabolites in 533 children • Age-specific, continuous reference intervals for plasma catecholamine metabolites • Dynamic reciprocal age-related increases in metanephrine and decreases in other metabolites • Reference intervals established and data made publically available • Biochemical testing using plasma metabolites now possible in young children [ABSTRACT FROM AUTHOR]

Phaeochromocytoma and extra adrenal paraganglioma are rare neuroendocrine tumours and have the potential to secrete adrenaline, noradrenaline and dopamine causing a myriad of clinical symptoms. Prompt diagnosis is essential for clinicians and requires a multidisciplinary specialist approach for the clinical and laboratory investigation, diagnosis, treatment and follow-up of patients. This paper is an integrated review of the clinical and laboratory evaluation and treatment of patients suspected to have phaeochromocytoma or paraganglioma, highlighting recent developments and best practices from recent published clinical guidelines. [ABSTRACT FROM AUTHOR]

We herein realize the first example of using a microporous HOF material HOF-BTB with high specific surface area for ratiometric fluorescent detection of pheochromocytomas and paragangliomas tumor biomarker 3-methoxytyramine (3-MT). HOF-BTB exhibits rapid, highly accurative and sensitive detection of 3-MT, with the detection limit as low as 46 ​nM. To the best of our knowledge, this HOF material achieves the best detection limit value among luminescent sensors, and is comparable to the detection limit of high-performance liquid chromatography or liquid chromatography-tandem mass spectrometry. [Display omitted] • A ratiometric fluorescence sensor based on the porous HOF-BTB material was reported. • HOF-BTB is the first report of HOFs for highly efficient sensing of biomarkers. • HOF-BTB achieves both superior accuracy and the best detection limit value. [ABSTRACT FROM AUTHOR]

Density functional theory calculations, with B3LYP functional and 6-311++G(d,p) basis set are performed with the aim to support the molecular structure and the spectroscopic characteristics of 3-methoxytyramine, the major extracellular metabolite of dopamine. Natural Bond Orbital (NBO) and Quantum Theory of Atoms in Molecules (QTAIM) analysis were used to explain the specific interactions in the most stable conformations in vacuum and water. The conformer resembling the crystallographic structure was compared to the experimentally available data and NMR spectra. The detailed vibrational spectral analysis and the assignments of the bands, done on the best-fit basis comparison of the experimentally obtained and theoretically calculated IR and Raman spectra, match quite well indicating DFT calculations as very accurate source of normal mode assignments. The obtained results demonstrate the applicability and performance of DFT calculations in conformational analysis of 3-methoxytyramine at the state of the isolated molecule. The molecular docking showed that the most stable conformation in vacuum was not the most stable one when docked in protein, proving that only the weak interactions stabilize the gaseous phase conformations. [ABSTRACT FROM AUTHOR]

Background Urinary dopamine, homovanillic acid and 4-hydroxy-3-methoxymandelic acid are established tests for diagnosis and monitoring of neuroblastic disease. We compared the diagnostic performance of total urinary 3-methoxytyramine, the O-methylated product of dopamine, to these three established tumour markers. Methods Urinary 3-methoxytyramine, dopamine, homovanillic acid and 4-hydroxy-3-methoxymandelic acid were measured by high-performance liquid chromatography with electrochemical detection on consecutive urine samples from histologically proven neuroblastic patients and controls. Patients with neuroblastic disease were further classified as untreated, advancing, residual or absent disease based on clinical and radiological criteria. Receiver operating characteristic curve analysis was used to compare the diagnostic performance of the four tumour markers. Results Urinary 3-methoxytyramine was well correlated with established tumour markers and its concentration correlated with disease activity. It was the most commonly elevated tumour marker in neuroblastic disease and showed similar sensitivity to dopamine and homovanillic acid. The diagnostic utility of urinary 3-methoxytyramine as measured by area under the receiver operating characteristic curve was similar to dopamine and homovanillic acid. Conclusion Our results support the use of urinary 3-methoxytyramine as a tumour marker in the diagnosis and the monitoring of neuroblastoma disease. [ABSTRACT FROM AUTHOR]

Among the urinary compounds measured in screening for pheochromocytomas and paragangliomas (PPGL), 3-methoxytyramine (3-MT) has a controversial place. In this study, we therefore aimed to assess the diagnostic value of an isolated elevated urinary 3-MT concentration. We collected clinical data from patients with an isolated elevated level of 3-MT/creatinine (3-MT/Cr) and from control subjects, without elevated 3-MT/Cr levels. Among 64 patients with an isolated elevation of 3-MT/Cr and 66 controls, 7 patients had PPGL compared to 8 controls. There was no difference in 3-MT/Cr values between the different types of tumor diagnosed. This study suggests that an elevated urinary 3-MT/Cr level is not specifically associated with dopamine-secreting PPGL. and may therefore have little value in PPGL screening. [ABSTRACT FROM AUTHOR]

Background: Plasma 3-methoxytyramine (3-MT), a metabolite of dopamine, is elevated in up to 28% of patients with head and neck paragangliomas (HNPGLs). As free dopamine is incorporated in circulating platelets, we determined dopamine concentration in platelets in patients with a HNPGL. Methods: A single center cohort study was performed between 2012 and 2014. Thirty-six patients with a HNPGL were compared to healthy controls (68 for dopamine in platelets and 120 for plasma 3-MT). Results: Dopamine concentration in platelets was elevated in HNPGL patients compared to healthy controls (median [interquartile ranges] 0.48 [0.32-0.82] pmol/109 platelets vs. 0.31 [0.24-0.47] pmol/109 platelets; p < 0.05), whereas plasma 3-MT concentration did not differ between both groups (0.06 [0.06-0.08] nmol/L vs. 0.06 [0.06-0.06] nmol/L; p = 0.119). Based on 68 healthy controls, the reference interval for dopamine concentration in platelets was 0.12-0.97 pmol/109 platelets. Six (16.7%) patients with a HNPGL demonstrated an increased dopamine concentration in platelets compared to three (8.3%) patients with an increased plasma 3-MT level (p = 0.053). The sensitivity and specificity were 16.7% and 98.5% for platelet dopamine and 8.3% and 97.5% for plasma 3-MT concentration (p = 0.37). Conclusions: Dopamine concentration in platelets is elevated in patients with a HNPGL compared to healthy subjects, and may be a novel biomarker for dopamine producing paraganglioma. [ABSTRACT FROM AUTHOR]

The activity of monoamine oxidases (MAOs) in the brain is often associated with neurodegenerative diseases. The study of MAOs in vivo or ex vivo is generally performed using MAO inhibitors and rarely using substrates. We present a pharmacological approach using intracerebral microdialysis to study the activity of MAO in the striatum and the prefrontal cortex of rats. It consists of applying ascending concentrations of 3-methoxytyramine (3-MT) as a substrate via the probes and measuring the indirect product homovanillic acid generated by MAO activity. We present herein the methodologies comprising our in-house stereotaxic procedures in rats, the microdialysis perfusion system and the substrate application, and the neurochemical analysis of the samples., (© 2023. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Background:Measurement of plasma metanephrine, normetanephrine and 3-methoxytyramine is useful in the diagnosis of phaeochromocytomas, but many assays require a large volume of plasma due to poor assay sensitivity, and often require lengthy sample preparation. Our aim was to develop a method for measurement of plasma metanephrines using a small sample volume with minimal hands-on preparation.Methods:Samples were deproteinised using 10K spin filters prior to online solid phase extraction using a Waters Acquity UPLC Online SPE Manager (Waters, Manchester, UK) coupled to a Waters Xevo TQ-S mass spectrometer(Waters, Manchester, UK). The assay was validated and results compared to a previously published method.Results:We achieved a limit of quantification of 37.5pmol/L for metanephrine and 3-methoxytyramine and 75pmol/Lfor normetanephrine using only 150mL of sample. The assay was linear up to 30,000pmol/L for all analytes and in a method comparison study results showed good agreement with a previously published LC-MS/MS assay. Conclusions: We have developed a simple method for measurement of plasma metanephrine, normetanephrine and 3-methoxytyramine using only 150mL of sample. There is minimal hands-on sample preparation required and the assay is suitable for routine use in a clinical laboratory. [ABSTRACT FROM PUBLISHER]

rapid and precise LC-ESI-MS-MS method for the parallel identification and quantification of dopamine, serotonin and their metabolites (homovanillic acid, 3-methoxytyramine, 3,4-dihydroxyphenylacetic acid and 5-hydroxyindolacetic acid) from rat brain tissue without any pre-analysis adjustment of the sample such as pre-concentration or derivatization has been developed. In particular, the reaction-monitoring mode was selected for its extremely high degree of selectivity and the stable-isotope-dilution assay for its high precision of quantification. Alternation the ionization polarity in the course of mass spectrometry detection enabled to determine substances susceptible to various ionization modes in only one analysis run. This fact, in combination with an easy pre-treatment step, constitutes the method straightforward and time undemanding. The developed method was characterized with a high precision (≤19.5%, determined as RSD), an acceptable accuracy (≥82.0%, determined as recovery), a low limit of detection (≤0.40 ng/100 mg brain tissue) and a low limit of quantification (≤0.42 ng/100 mg brain tissue). The method has been applied in a recent animal study. The levels of the studied neurotransmitters have been determined in the rat brain hippocampus, prefrontal cortex, and striatum in an animal model of schizophrenia induced by an acute dose of a dizocilpine. [ABSTRACT FROM AUTHOR]

Abstract: A sensitive assay method was developed for a parallel, rapid and precise determination of dopamine and its metabolites, homovanillic acid, 3-methoxytyramine and 3,4-dihydroxyphenylacetic acid, from brain microdialysates. The method consisted of a pre-treatment step, freeze-drying (lyophilization), to concentrate dopamine and its metabolites from the microdialysates, and a detection step using liquid chromatography combined with electrospray ionization tandem mass spectrometry (LC–ESI-MS/MS). In particular, the reaction monitoring mode was selected for its extremely high degree of selectivity and the stable-isotope-dilution assay for its high precision of quantification. The developed method was characterized by the following parameters: the precision of the developed method was determined as ≥88.6% for dopamine, ≥89.9% for homovanillic acid, ≥86.1% for 3-methoxytyramine and ≥88.1% for 3,4-dihydroxyphenylacetic acid; the mean accuracy was determined as ≥88.2% for dopamine, ≥88.3% for homovanillic acid, ≥85.9% for 3-methoxytyramine and ≥88.6% for 3,4-dihydroxyphenylacetic acid. The developed method was compared to (1) other combinations of pre-treatment methods (solid phase extraction and nitrogen stripping) with LC–MS and (2) another detection method, liquid chromatography, with electrochemical detection. The novel developed method using combination of lyophilization with LC–ESI-MS/MS was tested on real samples obtained from the nucleus accumbens of rat pups after an acute methamphetamine administration. It was proven that the developed assay could be applied to both a simultaneous analysis of all four substrates (dopamine, homovanillic acid, 3-methoxytyramine and 3,4-dihydroxyphenylacetic acid) in microdialysis samples acquired from the rat brain and the monitoring of their slight concentration changes on a picogram level over time following methamphetamine stimulus. [Copyright &y& Elsevier]

Levodopa and dopamine have been abused as performance-altering substances in horse racing. Urinary 3-methoxytyramine is used as an indicator of dopaminergic manipulation resulting from dopamine or levodopa administration and is prohibited with a urinary threshold of 4 µg mL−1 (free and conjugated). A simple liquid chromatographic (LC)/mass spectrometric (MS) (LCMS) method was developed and validated for the quantification and identification of 3-methoxytyramine in equine urine. Sample preparation involved enzymatic hydrolysis and protein precipitation. Hydrophilic interaction liquid chromatography (HILIC) was selected as a separation technique that allows effective retention of polar substances like 3-methoxytyramine and efficient separation from matrix compounds. Electrospray ionization (ESI) in positive mode with product ion scan mode was chosen for the detection of the analytes. Quantification of 3-methoxytyramine was performed with fragmentation at low collision energy, resulting in one product ion, while a second run at high collision energy was performed for confirmation (at least three abundant ions). Studies on matrix effects showed ion suppression depending on the horse urine used. To overcome the variability of the results originating from the matrix effects, isotopic labelled internal standard was used and linear regression calibration methodology was applied for the quantitative determination of the analyte. The tested linear range was 1-20 µg mL−1. The relative standard deviations of intra- and inter- assay analysis of 3-methoxytyramine in horse urine were lower than 4.2% and 3.2%, respectively. Overall accuracy (relative percentage error) was less than 6.2%. The method was applied to case samples, demonstrating simplicity, accuracy and selectivity. Copyright © 2009 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Abstract: The herbicide paraquat (PQ) is a strong redox agent that participates in the formation of reactive oxygen species (ROS) and induces toxicity in the nigrostriatal dopaminergic system. In this study, we investigated the effect of PQ on dopamine (DA) and glutathione levels in the substantia nigra (SN) of mice. Male C57BL/6 mice (aged 7 weeks and 23–25g) were used for this study. The mice were treated with normal saline (vehicle) and PQ (10mg/kg, i.p.) twice weekly for three consecutive weeks. We measured changes in tyrosine hydroxylase (TH) immunoreactivity, DA and its metabolites, and glutathione (reduced and oxidized) in the SN. After repeated PQ administration, the density of TH-positive neurons in the substantia nigra pars compacta (SNpc) decreased as compared to the control. Levels of DA and homovanillic acid (HVA) decreased significantly in the PQ-treated mice (p <0.05), but levels of 3,4-dihydroxyphenylacetic acid (DOPAC) and 3-methoxytyramine (3-MT) did not change. The rate of DA oxidation increased significantly in the SNpc, whereas the O-methylation pathway remained unchanged. Levels of reduced glutathione (GSH) in the SNpc decreased more in the PQ group than in the control group, while levels of oxidized glutathione (GSSG) increased in same region. We propose that repeated PQ injection induces dopaminergic neurotoxicity through generation of oxidative stress, and that this toxicity is related to the decline of GSH in the SNpc. The neurotoxic mechanism may specifically involve enhancement of the oxidative pathway of DA metabolism through coupling with the antioxidant GSH system of the SN. [Copyright &y& Elsevier]

Copyright of Revue Francophone des Laboratoires is the property of Elsevier B.V. and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

The lanthanide functionalized MOFs with an exceptional performance for early diagnosis of pheochromocytomas and paragangliomas was constructed as a multifunctional sensing platform based on postsynthetic modification strategy. [Display omitted] • It is the first report in designing a luminescent sensor to determine the biomarker 3-methoxytyramine. • The proposed sensor Eu3+@Al-MOF is applicable in the detection of 3-methoxytyramine in urine system with excellent performance. • The test paper creates a portable analysis tool for visual and selectivity detection of 3-methoxytyramine. As a main extracellular metabolite of dopamine, 3-methoxytyramine (3-MT) is considered a potential biomarker of pheochromocytomas and paragangliomas. Therefore, the determination of 3-MT is of great significance in the early diagnosis of disease. However, it remains challenging for detecting 3-MT in consideration of sensitivity and accuracy. Here, a luminescent Eu3+ functionalized metal-organic frameworks （Eu3+@Al-MOF）with ultra-high chemical stability was constructed based on postsynthetic modification. Such a rational design greatly enhances the fluorescence signal of Eu3+@Al-MOF and it is endowed with excellent properties as a luminescent sensor to detect 3-MT in urine system. Intriguingly, the strong red emitting derived from antenna effect was significantly interdicted upon addition of 3-MT through the interaction between 3-MT and the ligand. The proposed sensing system exhibited many appealing analytical performances, such as excellent selectivity, high sensitivity and quick response. Remarkably, the developed paper-based sensor not only provides a portable and reliable strategy for direct detection of 3-MT but also expands the application of visual analysis tools. This work represents the first effort in designing a luminescent sensor to determine the metabolite biomarker 3-MT level and provides a new method for biomedical analysis. [ABSTRACT FROM AUTHOR]

Chronic haloperidol treatment induces depolarization block in midbrain dopamine neuronal systems. We studied the effect of this treatment on nigrostriatal dopamine catabolism using microwave fixation in situ of the brain to prevent post-mortem changes. Male Sprague-Dawley rats were given haloperidol (0.4 mg/kg/day, i.p.) or vehicle for 21 days. On day 22, some rats in each group received a haloperidol challenge (0.4 mg/kg, i.p.), and the remaining rats were given the vehicle. Dopamine metabolite levels 60 min after the challenge were assayed by combined gas chromatography-mass fragmentography. Haloperidol pretreatment significantly modified haloperidol challenge effect on regional dopamine metabolite contents. The challenge elevated all striatal metabolites studied similarly in the chronic vehicle- or chronic haloperidol-pretreated rats. In contrast, it did not significantly affect nigral dopamine metabolites except it elevated 3,4-dihydroxyphenylacetic acid in the haloperidol-pretreated rats. A linear correlation between the nigral and striatal contents of 3-methoxytyramine (R = 0.72, p = 0.03), and a trend for correlation (R = 0.65, p = 0.06) between the respective 3,4-dihydroxyphenylacetic acid contents were found after the haloperidol challenge in the vehicle-pretreated rats only. These results suggest that chronic haloperidol treatment uncouples somatodendritic dopamine turnover and release from those in the axon terminals of nigrostriatal dopamine neurons. [ABSTRACT FROM AUTHOR]

The influence of sampling variables on the concentration of the dopamine metabolites 3-methoxytyramine (3MT), dihydroxyphenylacetic acid (DOPAC) and homovanilic acid (HVA) was examined in equine urine. A logarithmic transformation of the data for all horses gave distribution which approximated the normal distributions for each metabolite. The mean urinary concentration of 3MT in horses was 214ng/mL and the application of a threshold with a probability of 1 in 10,000 gave an actionable level of 4μg/mL. Environmental variables were not forensically significant in determining the population distribution. HVA was not found to be a reliable indicator of dopamine or levodopa administration. [Copyright &y& Elsevier]

In weaver mice, besides a postnatal cerebellar developmental anomaly probably caused by alterations of an inwardly rectifying K+ channel, there is a progressive loss of mesencephalic dopaminergic neurons. To further evaluate this deficit, endogenous dopamine and its metabolites were measured in 22 brain regions from heterozygous (wv/+) and homozygous (wv/wv) mutants, and compared to wild type (+/+) mice. In both wv/+ and wv/wv mutants there were profound dopamine depletions in all regions; these changes were accompanied by decreases in metabolites but with an increase of turnover indexes. Dopamine D1 and D2 receptors were examined by autoradiography, and their distribution was conserved. The results show that the dopaminergic deficit is widespread to all areas of innervation, and is probably compensated for by an increased turnover. Abnormal developmental growth signals, or aberrant cellular responses, may result in defective neurite formation of the midbrain dopaminergic neurons, leading to their postnatal death. [ABSTRACT FROM AUTHOR]

The effects of neuroleptics have been attributed to dopamine (DA) receptor blockade; however, other neurotransmitters, in particular serotonin (5-HT), have also been implicated. In this study, we examined the effects of clozapine and haloperidol on the distribution of DA and 5-HT transporters, on endogenous DA, 5-HT and their major metabolites, and on 5-HT1A receptors. Adult male Sprague-Dawley rats were treated with either haloperidol (1 mg/kg/day, i.p.), clozapine (20 mg/kg/day, i.p.) or saline for 21 days, and following 3 days of withdrawal, the brains were removed. Tissue levels of DA and 5-HT and their metabolites were measured by high-performance liquid chromatography in 16 brain regions, while quantitative autoradiography with [125I]RTI-121, [3H]citalopram and [3H]8-OH-DPAT was employed to label DA transporters, 5-HT transporters and 5-HT1A receptors, respectively. After haloperidol, densities of 5-HT transporters were increased in the dorsal insular cortex and in the ventral half of caudal neostriatum, while 5-HT1A receptors augmented in cingulate cortex but decreased in the entorhinal area. After clozapine, [3H]citalopram labelling was increased in ventral hippocampus, ventral caudal neostriatum and nucleus raphe dorsalis, but decreased in medio-dorsal and latero-dorsal neostriatum as well as in substantia nigra. Binding of [3H]8-OH-DPAT following clozapine was decreased in frontal, parietal, temporal and entorhinal cortices but increased in the CA3 division of Ammon's horn. The changes in 5-HT transporters in nucleus raphe dorsalis and substantia nigra, as well as the 5-HT1A receptor down-regulations caused by clozapine but not by haloperidol, may explain effects obtained with clozapine and other atypical neuroleptics. There were no modifications in densities of DA transporters, nor of tissue DA levels, after the chronic neuroleptic treatments. The results are in line with previous suggestions that a certain degree of tolerance to neuroleptics develops, in spite of profound D1 and D2 receptor changes that persist during the entire chronic treatment with these psychotropic agents. [ABSTRACT FROM AUTHOR]

The accumulation of 3-methoxytyramine (3-MT), the O-methylated metabolite of dopamine (DA), in rat striatum was used to assess the effects of drugs on dopaminergic activity. This was accomplished by pretreating rats with pargyline to completely inhibit 3-MT catabolism. Under the conditions used, 3-MT accumulation was linear over time for at least 90 minutes. Apomorphine and γ-butyrolactone, drugs which depress the activity of DA-containing neurons, decreased striatal 3-MT accumulation; whereas typical neuroleptics (haloperidol, fluphenazine, chlorpromazine), which increase the activity of DA-containing neurons, increased striatal 3-MT accumulation. In addition, a number of other drugs which block DA receptors and exert various atypical actions on dopaminergic functioning were examined. These 'atypical' compounds (clozapine, buspirone, molindone) also increased striatal 3-MT accumulation, but were generally less potent than the typical neuroleptics examined. Moreover, the potencies of the typical neuroleptics and 'atypical' compounds that were tested appear to be somewhat related to their affinities for D-2 DA receptors, as measured by their abilities to displace H-spiperone from rat striatal membrane preparations. Interestingly, this relationship was less evident when NaCl was omitted from the H-spiperone binding assay buffer. The potential antipsychotic drugs, BW 234U and SCH 23390, were also investigated for their effects on 3-MT accumulation and H-spiperone binding, and they were relatively inactive in both of these measures of dopaminergic activity. The observation that SCH 23390, a D-1 DA receptor antagonist, had negligible effects on 3-MT accumulation suggests that D-1 receptor blockade, unlike D-2 blockade, may have minimal effects on DA release. The relevance of the above findings to understanding the effects of drugs on dopaminergic transmission and the advantages of measuring 3-MT accumulation instead of basal 3-MT concentrations are discussed. [ABSTRACT FROM AUTHOR]

The value of 3-methoxytyramine (3-MT) as an indicator of impulse-related dopamine (DA) release has been assessed in rat corpus striatum. Moreover, the turnover of 3-MT was estimated by measuring its disappearance rate after COMT inhibition. Quantitation of 3-MT and DA was performed by gas chromatography/mass spectrometry (selected ion monitoring). Haloperidol in doses between 0.05 and 3 mg/kg p.o. did not increase endogenous 3-MT levels at any time up to 24 h after its administration, whereas it dose-dependently increased homovanillic acid and 3,4-dihydroxyphenylacetic acid. However, in doses above 0.1 mg/kg p.o., it enhanced the accumulation of 3-MT in clorgyline-pretreated animals. Conversely, baclofen in doses of 2 mg/kg i.p. and above decreased endogeneous 3-MT levels, but reduced the accumulation of this amine only poorly at 20 mg/kg i.p. in clorgyline-pretreated rats. A number of dopamine agonists, apomorphine (0.5 mg/kg i.v.), dipropyl-ADTN (0.03 mg/kg i.v.), but not bromocriptine (1 mg/kg i.v.) reduced endogenous 3-MT levels 10 min after administration by approximately 50%. The DA releasing agents d-amphetamine and methylphenidate showed different effects: the former increased endogenous 3-MT greatly, whereas the latter was without effect. The difference is likely to be related to the MAO inhibitory properties of amphetamine. 3-MT disappeared rapidly after COMT inhibition with 50 mg/kg i.v. tropolone (half-life of the initial disappearance about 1 min). The curve flattened off after 5-10 min. Turnover was calculated to be about 7 nmol/g/h, which corresponds to about a third of the turnover of DA. Our results suggest that an important part of DA metabolism occurs through 3-MT and that this amine is very effectively deaminated by MAO-A, so effectively indeed that increased formation does not increase its endogeneous levels. It appears, therefore, that 3-MT is not suitable as an indicator of increased DA release. However, it seems to have some value for an assessment of lowered DA release. [ABSTRACT FROM AUTHOR]

3-Methoxytyramine and dopamine accumulated in vivo in rat brain after monoamine oxidase inhibition with pargyline HCl. A dose of 100 mg/kg i.p. appeared to inhibit monoamine oxidase completely and led to a linear accumulation of 3-methoxytyramine for the first 90 min. Axotomy of the ascending dopaminergic fiber tract by means of a complete transverse cerebral hemisection almost completely blocked 3-methoxytyramine formation provided that catecholamine synthesis was inhibited with H 44/68 (methylester of α-methyl-p-tyrosine). The dopamine receptor agonist apomorphine, 0.5 mg/kg i.p., decreased, while the dopamine receptor antagonist haloperidol, 2 mg/kg i.p., accelerated 3-methoxytyramine formation. γ-Butyrolactone, 750 mg/kg i.p., not only decreased 3-methoxytyramine formation per se but also effectively antagonized the haloperidol-induced increase in 3-methoxytyramine accumulation. 3-Methoxytyramine formation after inhibition of monoamine oxidase appears to be a reliable indicator of impulse-induced dopamine release and metabolism. [ABSTRACT FROM AUTHOR]

Sibutramine hydrochloride, a novel monoamine reuptake inhibitor antidepressant, has been studied to determine whether it alters dopaminergic function in the brain. Its effects have been compared with bupropion, a dopamine reuptake inhibitor, and methamphetamine, a dopamine reuptake inhibitor and releasing agent. Sibutramine (0.1-3 mg/kg PO) and methamphetamine (0.3-30 mg/kg PO) both prevented reserpine (0.75 mg/kg IV) ptosis in rats with ED values of 0.6 mg/kg and 4.2 mg/kg, respectively. Bupropion (10-100 mg/kg PO) was ineffective against reserpine ptosis. The efflux of [H]-dopamine from preloaded rat striatal slices was not altered by 10-10 M concentrations of sibutramine, BTS 54 354, BTS 54 505 (secondary and primary amine metabolites, respectively) or bupropion. In contrast, methamphetamine (10-10 M) caused a significant concentration-dependent increase in [H]-dopamine release. Sibutramine (3 mg/kg IP or 6 mg/kg PO) and bupropion (10 mg/kg IP or 430 mg/kg PO) did not alter 3-methoxy-tyramine (3-MT) levels in rat striatum. Striatal 3-MT concentrations were, however, dose-dependently increased by methamphetamine (0.3-10 mg/kg IP or 0.42-4.2 mg/kg PO). Sibutramine (6 mg/kg PO) did not induce circling in rats with unilateral 6-hydroxydopamine lesions of the nigrostriatal dopaminergic neuronal tract. Bupropion (10-100 mg/kg PO) did not induce circling at the lowest dose, but caused increasing ipsilateral rotation at higher doses. Methamphetamine (0.42 or 4.2 mg/kg PO) induced ipsilateral circling with marked effects at the higher dose. In a two-choice lever pressing model using rats trained to discriminate d-amphetamine (0.5 mg/kg IP) from saline, sibutramine (0.3-3 mg/kg IP) generalised to the saline lever. Bupropion (3-30 mg/kg IP) generalised to d-amphetamine at the highest dose, while methamphetamine (0.1-5 mg/kg IP) generalised to this lever at doses as low as 0.3 mg/kg. Overall, the rank order of potency for enhancing central dopaminergic function is methamphetamine > bupropion >> sibutramine. The data therefore indicate that dopamine is unlikely to be an important pharmacological target for reuptake inhibition by sibutramine. [ABSTRACT FROM AUTHOR]

Antipsychotic medications appear to exert their therapeutic effects by blocking D2 receptors. While D2 blockade occurs rapidly, reduction in psychotic symptoms is often delayed. This time discrepancy has been attributed to the relatively slow development of depolarization inactivation (DI) of dopaminergic neurons. The reduced firing rates associated with DI has been hypothesized to reduce dopamine release and thus psychotic symptoms. Studies assessing changes in dopamine release during chronic neuroleptic treatment, using microdialysis and voltammetry, have been inconsistent. This may be due to methodological differences between studies, the invasive nature of these procedures, or other confounds. To investigate the effects of DI on dopamine release, 3-MT accumulation, an index of dopamine release that does not involve disruption of brain tissue, was measured during acute and chronic neuroleptic treatment. These results are compared with those using other techniques. 3-MT levels remained elevated after chronic treatment, suggesting that DI does not markedly reduce release. Regulation of dopamine release during DI was examined using two techniques known to block dopamine neuronal impulse flow. 3-MT levels were markedly reduced by both, implying that DI does not alter the portion of dopamine release mediated by neuronal impulse flow. Overall, studies to date suggest that the delayed therapeutic effects of neuroleptics are not due to reductions in impulse dependent dopamine release. Recent studies using a neurodevelopmental animal model of schizophrenia have pointed to altered pre- and post-synaptic indices of dopamine neurotransmission. The results suggest that neuroleptics may exert their therapeutic effects, in part, by limiting the fluctuations in dopamine release, and raise new issues for future research. [ABSTRACT FROM AUTHOR]

The acute effects of N-methyl-4-phenyl-l, 2, 3, 6-tetrahydropyridine (NMPTP) on mouse central monoaminergic neurons were investigated. In striatum, two hours after the injection of NMPTP (50 mg/kg s.c.) there was a seven-fold increase in 3-methoxytyramine that coincided with a 55 % decrease in dopamine. In addition, the same dose of NMPTP caused an acute increase in mouse motor activity; this effect was antagonized by haloperidol or by pretreatment with reserpine. These data indicate that NMPTP initially causes a pronounced release of dopamine into the extraneuronal space. [ABSTRACT FROM AUTHOR]

Intraperitoneal administration of pargyline HCl induced a dose-dependent accumulation of 3-methoxytyramine and normetanephrine in mouse brain in vivo. As judged by the decrease of 5-hydroxyindole acetic acid levels a dose of 200 mg/kg of pargyline appeared to inhibit monoamine oxidase completely. This dose led to an approximately linear accumulation of 3-methoxytyramine and normetanephrine during the first 3 hours. γ-Butyrolactone, 750 mg/kg i.p. reduced the accumulation of 3-methoxytyramine despite a marked increase of dopamine. (+)-Amphetamine stimulated 3-methoxytyramine as well as normetanephrine accumulation at doses of 3 and 10 mg/kg i.p. In line with the concept of receptor-mediated negative feedback control of catecholaminergic transmission the dopamine receptor agonists apomorphine, 0.3 mg/kg i.p., lisuride, 0.05-0.3 mg/kg i.p., and bromocriptine, 10 mg/kg i.p., decrease 3-methoxytyramine formation while the dopamine receptor blocking agent haloperidol, 1 mg/kg i.p., led to a 3-fold increase. The α-adrenoceptor agonist clonidine, 0.1 mg/kg i.p., reduced the formation of normetanephrine and the α-adrenoceptor antagonists yohimbine, 10 mg per kg i.p., phenoxybenzamine, 20 mg/kg i.p., and mianserine, 50 mg/kg i.p., stimulated normetanephrine accumulation 1.5- to 4-fold. 3-Methoxytyramine and normetanephrine accumulating after inhibition of monoamine oxidase appear to be reliable indicators of dopamine and noradrenaline release and metabolism. [ABSTRACT FROM AUTHOR]

A method for the chromatographic separation (Dowex 50 W, X-4) and fluorimetric determination of 3-methoxytyramine in brain tissue is described. The chromatographic procedure is an extension of the method for the separation of noradrenaline, dopamine and 5-hydroxytryptamine published by Atack and Magnusson (1970) and can be combined with that for monoamine precursors described by Kehr et al. (1972). 3-Methoxytyramine is eluted in N ethanolic (50%)-HCl and oxidized with potassium ferricyanide in ammonium hydroxide solution. Compared with previously published methods, the sensitivity is increased 2 to 5 fold. Reproducibility, recovery and specificity of the method are satisfactory. The level of 3-methoxytyramine found in normal rat brain is below the limit of sensitivity (10 ng/g). The amount of 3-methoxytyramine accumulating after inhibition of monoamine oxidase (MAO) differs depending on the inhibitor used. These differences in 3-methoxytyramine formation might be due to incomplete MAO inhibition, changes in dopamine synthesis, and releasing activity of the compounds investigated. [ABSTRACT FROM AUTHOR]

Dementia is exacerbated by loss of appetite and amotivation, and recent studies have indicated that ninjin'yoeito improves anorexia and amotivation. Previous studies suggest that ninjin'yoeito inhibits dopamine-metabolizing enzymes and enhances dopamine signaling. However, whether ninjin'yoeito increases dopamine content in living cells remains unclear. Here, PC12 cells were used to examine whether ninjin'yoeito affects the dopamine metabolic pathway. Dopamine content significantly increased 3 h after treatment ninjin'yoeito extract. Concomitantly, the levels of 3-methoxytyramine and 3,4-dihydroxyphenylacetic acid were significantly reduced. The effects of components of ninjin'yoeito on the dopamine metabolic pathway were also assessed. Treatment with onjisaponin B, nobiletin, and schisandrin, and the ingredients of Polygalae Radix, Citri Unshiu Pericarpium, and Schisandrae Fructus increased dopamine content and decreased its metabolite content in the culture media. Our findings suggest that ninjin'yoeito improves anorexia and amotivation by inhibiting metabolic enzyme and increasing the dopamine content in cells., (© The Author(s) 2021. Published by Oxford University Press on behalf of Japan Society for Bioscience, Biotechnology, and Agrochemistry.)