Mice striatal samples were homogenized with 0.1 M perchloric acid (1 mg tissues in 100 μL perchloric acid) and 0.1 mM EDTA, treated by ultrasonic and centrifuged at 20,000 rpm for 30 min as reported [27 ]. Then supernatant liquids were collected for measuring. The mobile phase is a mixed solution consisting of 90 mM sodium phosphate monobasic, 1.7 mM 1-octanesulfonic acid, 50 mM citrate, 50 μM EDTA, 10% acetonitrile with the flow rate of 0.2 ml/min. In parallel, for quantitative standard curve calculation, stock standards solution for dopamine and dihydroxy-phenyl aceticacid (DOPAC) (Sigma-Aldrich, USA) were prepared in 0.1 M HClO4. Amount of 10 μl of prepared supernatant or standard solution was injected into the mobile phase and tested by ESA Coulochem III electrochemical detector (Coulochem III, Thermo Fisher Scientific). Samples were measured and peaks were quantified. The concentrations of monoamines were quantified by comparing with the standard using ClarityChrom software (Knauer, Germany) and then the contents in stratum were converted according to the dilution ratio. The standard curve of DA and DOPAC were shown in Supplementary Fig. 1a and b .
Claritychrom software
Manufactured by Knauer
Sourced in Germany
ClarityChrom is a software solution designed for the control and data analysis of liquid chromatography systems. It provides a comprehensive platform for managing the operation of chromatographic instruments, as well as for processing and interpreting the acquired data.
Automatically generated - may contain errors
Lab products found in correlation
Esa coulochem 3 electrochemical detector, by Thermo Fisher Scientific (1 mentions)
Methanol, by Merck Group (1 mentions)
Ethylenediaminetetraacetic acid edta, by Merck Group (1 mentions)
Nucleosil c18 column, by Macherey-Nagel (1 mentions)
Whatman, by Cytiva (1 mentions)
Spherisorb ods 2 column, by Merck Group (1 mentions)
Smartline, by Knauer (1 mentions)
Smartline autosampler 3800, by Knauer (1 mentions)
4 protocols using claritychrom software
1
Striatal Monoamine Quantification Protocol
2
Quantification of Dopamine and Metabolites
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The concentrations of DA and the corresponding metabolites (3,4-dihydroxyphenylacetic acid [DOPAC] and homovanillic acid [HVA]) were evaluated by HPLC with electrochemical detection (potential set at 0.8 V with respect to Ag/AgCl reference electrode) and a glassy carbon electrode. Samples were homogenized in ice-cold 0.1 M HClO4 solution and centrifuged (13,000 x g, 15 min) to precipitate proteins. The supernatant was filtered (0.2 μm pore size; Whatman, USA) and a 20 μl aliquot was injected onto the Nucleosil C-18 column, 250 mm, 5 μm particle size (Macherey–Nagel, Germany). The mobile phase was 32 mM sodium phosphate, 34 nM citric acid, 1 mM octanesulfonic acid, and 27 μM ethylenediaminetetraacetic acid (EDTA) (Sigma, USA) in deionized (18.3 mΩ) water and 12% methanol (Merck, Germany). The monoamines were separated using a 0.8 ml/min flow rate. Samples were quantified by comparing with standard (Sigma, USA) solutions (external calibration) using ClarityChrom software (Knauer, Germany).
3
HPLEC Analysis of Ammonium Formate Mixtures
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All experiments were performed at the temperature of 25 °C, applying 100 bar pressure to the cushion pressurizing the adsorbent layer. The mobile phase was the mixture of water/methanol (3/2 v/v) with the addition of 80 mM ammonium formate (final concentration), pH 3.0 (obtained by addition of formic acid to ammonium formate solution in water). Most of the HPLEC equipment modules were controlled by a computer with Clarity Chrom software (Knauer, Berlin, Germany). Only the high-voltage power supply, the pressure supply, and the circular thermostat were programed independently.
4
HPLC Analysis of Phenolic Compounds in Tree Leaves
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Chromatographic separation was performed as previously reported by Vinha et al. (2002) (link), with an analytical HPLC unit (Knauer Smartline), equipped with a Knauer Smartline autosampler 3800, and a Knauer Diode Array Detector (DAD). A reversed-phase Spherisorb ODS2 column was used during analysis (250 x 4.6 mm, 5 μm particle size, Merck, Darmstadt, Germany). The used solvent system was a gradient of water–formic acid (19:1) and methanol, applied at a flow rate of 0.9 mL min−1. Spectral data from all peaks were accumulated within the 200–400 nm range. Chromatograms were recorded at 280, 320 and 350 nm, and data were managed on ClarityChrom® software (Knauer, Berlin, Germany). Phenolic compounds were quantified through the comparison performed with known amounts of external standards: hydroxytyrosol, oleuropein, chlorogenic acid and rutin were quantified at 280 nm, caffeic acid at 320 nm, and verbascoside, apigenin-7-O-glucoside, luteolin-7-O-glucoside and luteolin at 350 nm. HPLC analyses were performed using two technical replicates for each extract. The means of the four replicates for each tree leaf sample were then calculated. Phenolic compounds were expressed as the amount of phenolics per dry weight (DW) of leaf extract (mg/g of DW).
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