The determination of norepinephrine and epinephrine in plasma by HPLC with amperometric detection was modified, giving detection limits of 25 ng/1 and 18 ng/1 for norepinephrine and epinephrine, respectively, using l ml plasma. In order to achieve this sensitivity,* it was necessary to minimize the background noise by modification of Instrumentation and specimen handling. Particularly important was the extra purification of the reagents, the application of micro-bore HPLC, the enzymatic cleavage of uric acid and temperature control of the amperometric cell and the amplifier. Comparison of the present method with the radioenzymatic determination of catecholamines resulted in coefficients of correlation of r = 0.924 and 0.919 for norepinephrine and epinephrine, resp. (n = 38). The concentrations of the 38 different samples used for the comparison were in the physiological ränge. Bestimmung der Katecholamine im Plasma durch HPLC und amperometrische Detektion -Vergleich mit einer radioenzymatischen MethodeZusammenfassung: Die Bestimmung von Noradrenalin und Adrenalin im Plasma mit HPLC und amperometrischer Detektion wurde so modifiziert, daß -bei einem Probenvolumen von l ml Plasma -eine Nachweisgrenze von 25 ng/1 für Noradrenalin und von 18 ng/1 für Adrenalin erreicht würde. Diese Empfindlichkeit konnte durch Erniedrigung des Untergrundrauschens erreicht werden, indem die Meßapparatur und die Probenaufarbeitung modifiziert wurden. Besonders wichtig sind hierbei die Reinigung der verwendeten Reagenzien, die Verwendung von "micro-bore"-Säulen, die enzymatische Spaltung der Harnsäure sowie die Temperierung der amperometrischen Zelle und des Verstärkers. Bei dem Vergleich der hier beschriebenen Methode mit der radioenzymatischen Methode ergaben sich Korrelationskoeffizienten von r = 0,924 und 0,919 für Noradrenalin und Adrenalin (n = 38), wobei die Katecholaminkonzentrationen in allen Proben im physiologischen Bereich waren.
The concentration of free and conjugated norepinephrine (NE), epinephrine (E) and dopamine (DA) were measured by a modified radio-enzymatic assay in the plasma and in the cerebrospinal fluid (CSF) of 45 patients with normal and in 21 patients with disturbed blood-CSF barriers. In patients with an undisturbed blood-CSF barrier the free NE and E in CSF were 128 +/- 45 ng/l and 27 +/- 20 ng/l (mean values +/- S.E.), respectively, and represented about 50% of the average plasma values. Mean DA was not different in plasma (47 +/- 22 ng/l) and in CSF (41 +/- 19 ng/l). Both in plasma and in CSF, considerable higher free catecholamine (CA) levels were measured in patients with dysfunction of the blood-CSF barrier. In one patient with bacterial meningitis twofold higher concentrations of free NE and DA in CSF as compared with plasma were detectable. Sulfate conjugates of catecholamines are predominant in plasma and CSF. The contribution of conjugated CA to total CA in plasma from patients with normal blood-CSF barrier averaged 69.7%, 63.1% and 98.1% for NE, E and DA, respectively and was significantly lower in the CSF (p less than 0.001). In patients with disturbed blood-CSF barrier, the increases of conjugated CA were more pronounced in CSF than in plasma. Further, the contribution of conjugated NE and E to total NE and E in CSF was not only increased in patients with bacterial meningitis, but also in patients with renal insufficiency compared to the "control" patients (p less than 0.02 and p less than 0.001 resp.). Free and conjugated NE, E and DA in the plasma and CSF were related significantly (p less than 0.01 resp.) with stronger correlation for conjugated CA (p less than 0.001 resp.). These results together with findings in the literature, suggest that there is little or no rostral-caudal gradient in CSF CA conjugate concentrations and that even in patients with intact blood-CSF barrier plasma conjugated CA concentrations influence those in CSF. Thus only free CA levels in CSF may reflect the central adrenergic activity.
Bauersfeld, Diener, Knoll, Ratge and Wisser: Determination of urinary vanilmandelic acid and homovanillic acid 217 Summary: A method was developed for the simultaneous determination of urinary vanilmandelic acid and homovanillic acid, which included a two step prepurifteation and a reversed-phase high-performance liquid chromatography with amperometric detection. Conditions were evaluated for performing measurements with the amperometric detector free from electric interference.The method was linear between 2.5 and 100 / vanilmandelic acid and homovanillic acid with good precision (CV always less than 10%).The correlation between the present determination of vanilmandelic aciä and the procedure ofPisano et al. ((1962) Clin. Chim. Acta 7, 285^291) was very göod (r = 0.931). No interfering substances could be detected. Bestimmung von Vanillinmandelsäure und Homövanillinsäure im Urin mit Hochdruckflüssigkeitschromatographie und aniperometrischer DetektionZusammenfassung: Eine neue Methode zur gleichzeitigen Bestimmung von Vanillinmandelsäure und Homovanillinsäure im Urin wurde erarbeitet. Sie basiert auf der Kombination von HPLC und amperometrischer Detektion sowie einer zweistufigen Vortrennung. Es wurden die Versuchsbedingungen untersucht, die eine störungsfreie Messung mit dem amperometrischen Detektor ermöglichen. Das Verfahren ist von 2,5 bis 100 /imol/1 Vanillinmandelsäure und Hpmovanillinsäure linear, wobei der Variationskoeffizjent über den gesamten Bereich kleiner als 10% ist. Die Korrelation der Bestimmung der Vanillinmandelsäure mit der Methode vonPisano et al. ((1962) Clin. Chim. Actä 7, 285-291) ist hoch (r = 0,931). Störende Substanzen konnten nicht festgestellt werden.
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