Simple, sensitive assay of polyamines by high-performance liquid chromatography with electrochemical detection after post-column reaction with immobilized polyamine oxidase.

1989 ◽  
Vol 35 (8) ◽  
pp. 1694-1696 ◽  
Author(s):  
K Maruta ◽  
R Teradaira ◽  
N Watanabe ◽  
T Nagatsu ◽  
M Asano ◽  
...  
Keyword(s):  
Hydrogen Peroxide ◽  
Electrochemical Detection ◽  
High Performance ◽  
Immobilized Enzyme ◽  
Platinum Electrode ◽  
Reversed Phase ◽  
Polyamine Oxidase ◽  
Soybean Seedlings ◽  
Liquid Chromatographic ◽  
Urinary Polyamines

Abstract This simple, rapid liquid-chromatographic assay of urinary polyamines (putrescine, spermidine, spermine, and cadaverine) involves electrochemical detection with a post-column immobilized enzyme, polyamine oxidase (EC 1.4.3.6) from soybean seedlings. Polyamines are separated by isocratic ion-pairing reversed-phase chromatography, then enzymatically converted, with release of hydrogen peroxide, via the post-column reactor with immobilized polyamine oxidase; the hydrogen peroxide is detected by electrochemical oxidation on a platinum electrode. The detection limits for injected putrescine, spermidine, and spermine were 0.3, 0.5, 0.6, and 4 pmol, respectively, with linear ranges of two to three orders of magnitude. Reproducibility was also good, with CV values less than 7%. The efficiency of the immobilized enzyme column was not decreased after analysis of 300 urine samples. Putrescine and spermidine excretion in urine from patients with blood cancers and solid cancers was significantly increased.

Liquid-chromatographic assay of urinary 5-hydroxy-3-indoleacetic acid, with electrochemical detection.

1980 ◽  
Vol 26 (7) ◽  
pp. 907-909 ◽  
Author(s):  
Z K Shihabi ◽  
J Scaro
Keyword(s):  
High Performance Liquid Chromatography ◽  
Liquid Chromatography ◽  
Electrochemical Detection ◽  
High Performance ◽  
Indoleacetic Acid ◽  
Reversed Phase ◽  
Solvent Mixtures ◽  
Liquid Chromatographic ◽  
Phase Column ◽  
Reversed Phase Column

Abstract After extraction with two organic solvent mixtures, urinary 5-hydroxy-3-indoleacetic acid can be assayed by "high-performance" liquid chromatography on a reversed-phase column, with electrochemical detection. Compared to the nitrosonaphthol method (J. Biol. Chem. 216: 499, 1955), this method is more specific for detection of patients with carcinoid tumors.

Liquid-chromatographic assay of urinary 5-hydroxy-3-indoleacetic acid, with electrochemical detection.

1980 ◽  
Vol 26 (7) ◽  
pp. 907-909
Author(s):  
Z K Shihabi ◽  
J Scaro
Keyword(s):  
High Performance Liquid Chromatography ◽  
Liquid Chromatography ◽  
Electrochemical Detection ◽  
High Performance ◽  
Indoleacetic Acid ◽  
Reversed Phase ◽  
Solvent Mixtures ◽  
Liquid Chromatographic ◽  
Phase Column ◽  
Reversed Phase Column

Abstract After extraction with two organic solvent mixtures, urinary 5-hydroxy-3-indoleacetic acid can be assayed by "high-performance" liquid chromatography on a reversed-phase column, with electrochemical detection. Compared to the nitrosonaphthol method (J. Biol. Chem. 216: 499, 1955), this method is more specific for detection of patients with carcinoid tumors.

On-line cleavage of urinary estriol conjugates with immobilized beta-glucuronidase before liquid-chromatographic analysis.

1981 ◽  
Vol 27 (9) ◽  
pp. 1554-1557 ◽  
Author(s):  
L D Bowers ◽  
P R Johnson
Keyword(s):  
High Performance ◽  
Immobilized Enzyme ◽  
Gradient Elution ◽  
Storage Conditions ◽  
Reversed Phase ◽  
Liquid Chromatograph ◽  
On Line ◽  
Liquid Chromatographic Analysis ◽  
The Stability ◽  
Liquid Chromatographic

Abstract Urinary estriol measurement has been widely accepted as a useful indicator of fetoplacental status. Classically, glucuronide conjugates of estriol have been cleaved with soluble beta-glucuronidase (EC 3.2.1.31) before extraction and measurement. We have developed a system in which urine is injected directly into a "high-performance" liquid chromatograph and the conjugates are cleaved on-line in an immobilized beta-glucuronidase reactor. Estriol is quantitated with fluorescence detection after separation from other interfering species with a reversed-phase column. The stability of the immobilized enzyme under storage conditions and in the presence of the mobile phase are discussed. Only 150 mL/L methanol could be pumped through the reactor and onto the analytical column, but this allowed on-column preconcentration of the free estriol produced. Gradient elution bypassing the immobilized enzyme reactor eluted the compounds of interest without damaging the enzyme. Comparison with radioimmunoassay results yielded a slope of 0.97 (r = 0.996, n = 19).

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