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KMID : 0377619830450040243
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Korean Jungang Medical Journal
1983 Volume.45 No. 4 p.243 ~ p.254
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Biological Studies on Schizophrenia-A Rapid Microassay of Catecholamines by High Performance Liquid Chromatography-Radioenzymatic Method
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Abstract
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A direct rapid radioenzymatic assay in combination with high performance liquid chromatography (HPLC) for measuring simultaneously picogram quantities of nore pinephrine, epinephrine and dopamine has been developed, and the relationship of changes in cerebrospinal fluid (CSF) norepinephrine concentration during haloperidol treatment to changes in psychosis rating of twelve schizophrenic patients has been investigated.
The three catecholamines in 20?l of CSF are converted to their 0-methylated ana-logues by catechol-0-methyltransferase in the presence of S-adenosyl-L-1{methyl-3H} methionine. After the addition of tetraphenylborate, these derivatives are extracted. with diethylether at pH 8.2 followed by re-extraction with 0.1N HCI and then isol-ated by HPLC using, ?Bondapak C18column and 0.17 M acetic acid-60% methanol- 0.02 M Na2SO4 as solvent. Manual collection of each fraction is started two seconds after in appeared mark. The HPLC fraction of norepinephrine and epinephrine deri-vatives are oxidized to vanillin followed by extracting the vanillin with toluene, and dopamine derivatives fraction is extracted with toluene/isoamylalcohol and then re-extracted with 0.1N HCl.
Internal standard curve shows a good linearity from 5 to 30 picogram per samples, and incorporated radioactivity of norepinephrine, epinephrine and dopamine derivatives are 163±11.4, 125±10.5 and 112±3.9cpm per picogram and calculated sensitivities are 0.4, 0.3 and 1 picogram, respectively.
3H-Normetanephrine, 3H-metanephrine and 3H-methoxytytamine are well separated by HPLC, and each retention times are 3.52, 4.08 and 5.62 minutes and yields are 75.6?0.9, 76.1?0.8 and 70.1?0.4%.
Separation of the methylated derivatives by HLPC are performed within 8 minutes, and thus enabled us to complete the differential analysis of 15 samples (30 tubes) in a day.
CSF norepinephrine concentration in schizophrenic patients show rostral-caudal gradient and haloperidol induced decreases of norepinephrine and the decreases correlated significantly with psychosis rating,
From the above rsults, it is suggested that this method is more excellent than any others in sensitivity, accuracy, reproducibility and specificity of the differential assay, and CSF norepinephrine can be used for evaluation of neuroleptics treatment of schizophrenia by this method.
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