[Detection of monoamine neurotransmitters and its metabolites by high performance liquid chromatograph after pre-column derivatization of dansyl chloride column].
OBJECTIVE
To develop a high performance liquid chromatography (HPLC) for detection of monoamine neurotransmitters and its metabolites after pre-column derivatization with dansyl chloride.


METHODS
The C(18) chromatograph column (150 mm×4.6 mm×5 µm) was selected for detection, and derived by dansyl chloride (10 mg/ml) under the condition of 50°C water bath by pH11 buffer solution. 20 µl acetic acid acetone solution (1.0 mol/L) was then mixed in for termination of the reaction. Then the solution was cooling to room temperature, 0.1 mol/L acetic acid zinc-acetonitrile-tetrahydrofuran solution was adopted for mobile phrase, with the volume ratio at 62:35:3. The flow rate was 1.0 ml/min between 0-10 min, 2.0 ml/min between 10-35 min. The ultraviolet detection wavelength was 286 nm. The above method separately detected monoamine neurotransmitters and its metabolites and evaluated the limit of detection, accurate degree and accuracy degree.


RESULTS
The linear relations between each component was good in the range of 1 - 20 µg/ml (r = 0.999). The lowest detection limit of norepinephrine, dopamine, 5-hydroxytryptamine and the metabolites 3-methoxy-4-benzoglycols, homovanillic acid and 5-heteroauxin were separately 0.60, 0.80, 0.41, 0.21, 0.19 and 0.1 µg/ml; while the average recovery rates were between 78.5% - 95.9%, and the relative standard deviation (RSD) was 6.62%, 7.64%, 2.98%, 3.60%, 5.09% and 3.09%, respectively. In the process of selection and optimization of the chromatographic conditions, we observed the importance of metal ions to discretion, and discussed the temperature, pH of the buffer solution and dosage of dansyl chloride in derivation. Under the above conditions, the reaction was perfect, and the baseline of the detected materials thoroughly separated.


CONCLUSIONS
The method to detect monoamine neurotransmitters and its metabolites by HPLC and pre-column derivatization with dansyl chloride was established; and this method could provide reference for the detection of polyamine by HPLC.
Department of Preventive Medicine and Medical Examination, Xiangnan University, Chenzhou 423000, China. huangxiao1998@163.com
Xiao
X
Huang
Jia-wen
JW
Chen
Li-ping
LP
He
Xue-jun
XJ
Kang
D015306:Biogenic Monoamines / Q000032:analysis* / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D003619:Dansyl Compounds; D018377:Neurotransmitter Agents / Q000032:analysis* / Q000378:metabolism
D015306:Biogenic Monoamines; D003619:Dansyl Compounds; D018377:Neurotransmitter Agents; C029038:dansyl chloride