Measuring levels of biogenic amines and their metabolites in rat brain tissue using high-performance liquid chromatography with photodiode array detection

Abstract

We developed a method to detect biogenic amines and their metabolites in rat brain tissue using simultaneous high-performance liquid chromatography and a photodiode array detection. Measurements were made using a Hypersil Gold C-18 column (250 × 2.1 mm, 5 µm). The mobile phase was 5 mM perchloric acid containing 5 % acetonitrile. The correlation coefficient was 0.9995-0.9999. LODs (S/N = 3) and LOQs (S/N = 10) were as follows: dopamine 0.4 and 1.3 pg, 3, 4-dihydroxyphenylacetic acid 8.4 and 28.0 pg, serotonin 0.4 and 1.3 pg, 5-hydroxyindolacetic acid 3.4 and 11.3 pg, and homovanillic acid 8.4 and 28.0 pg. This method does not require derivatization steps, and is more sensitive than the widely used HPLC-UV method.

Comparison of Three Extraction Approaches for the Isolation of Neurotransmitters from Rat Brain Samples

“…The idea of the presented study was to discuss the most popular sample preparation approaches for the isolation of NTs, their precursor amino acids, and their metabolites from brain slices. To clearly demonstrate the pros and cons of the chosen analytical approaches, 200 mg of brain tissue sample was processed by mechanical homogenization with the most popular solvents (formic acid (FA) or perchloric acid (PCA) solutions) [ 11 , 12 , 13 , 14 , 15 ]. For the stability of the analytes, the homogenization was performed in a hand-made ice bath, and the entire experimental procedure was carried out in the dark.…”

CRISPR/sgRNA-directed synergistic activation mediator (SAM) as a therapeutic tool for Parkinson´s disease

“…Brain slices, 100 µm thick, were cut using a Leica microtome (RM2125 RTS) in ice-cold Krebs-Henseleit solution. Immediately, left and right striata sections were dissected and collected in an amber 1.5 ml Eppendorf tube with sterile PBS solution at 4 °C (100 µl) and 50 µl of perchloric acid 0.1 N, the tissue was manually homogenized, sonicated and ultracentrifuged, the supernatant was filtered through a 0.22 µm filter (Millipore GSWP04700) to assay for DA and DOPAC [ 31 , 32 ]. The medium culture samples were collected in an amber 1.5 ml Eppendorf tube with 50 µl of perchloric acid 0.1 N per 950 µl of medium, samples were shaken with a vortex for a minute and ultracentrifuged, the supernatant was filtered through a 0.22 µm filter (Millipore GSWP04700) to assay for DA and DOPAC.…”

Role of caloric vestibular stimulation in improvement of motor symptoms and inhibition of neuronal degeneration in rotenone model of Parkinson's disease – An experimental study

“…The collected supernatant was again centrifuged at 12,000g, 4 8C for 20 min and filtered through a 0.22 mm membrane filter. The collected filtrate (1 mL) was added into a sterile dark container and stored at À80 8C [18].…”

“…Dopamine contents were estimated by LC-20AP HPLC (Shimadzu Corporation, Kyoto, Japan) with photodiode array detector (PDA) detector as adopted by Gu et al [18]. The system was equipped with degassing unit, column oven, low pressure gradient and isocratic unit, LC-20AP Quaternary low pressure mixing pump and Shimadzu SPD-M20A UV-DAD detector with thermostated flow cell.…”