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PubMed1991

Correlations between polyamine analogue-induced increases in spermidine/spermine N1-acetyltransferase activity, polyamine pool depletion, and growth inhibition in human melanoma cell lines.

The polyamine analogue, N1,N12-bis(ethyl(-spermine (BESPM), is known to suppress ornithine and S-adenosylmethionine decarboxylase levels, deplete intracellular polyamine pools, and inhibit cell growth. Among human melanoma cell lines, MALME-3 cells were found to be typically sensitive to the antiproliferative activity of the BESPM, whereas LOX cells were atypically insensitive to the analogue. A comparison of polyamine-related parameters revealed that the most differentially altered activity between the 2 BESPM-treated cell lines was that of spermidine/spermine N1-acetyltransferase (SSAT), which increased from 50 pmol/min/mg to greater than 10,000 pmol/min/mg in MALME-3 cells and from 16 pmol/min/mg to only 120 pmol/min/mg in LOX cells over 48 h. The basis for the large difference seems to be related to increased enzyme synthesis in both cell lines coupled with differences in prolongation of SSAT half-life (greater than 12 h in MALME-3 cells versus 1.6 h in LOX cells) after BESPM treatment. In MALME-3 cells, SSAT accumulation was found to be differentially modulated by the BESPM homologues, N1,N11-bis-(ethyl)norspermine and N1,N14-bis-(ethyl)homospermine, which were 5-fold more and 9-fold less effective, respectively, than BESPM in increasing SSAT but similar in analogue uptake and effects on polyamine biosynthesis and cell growth inhibition. Treatment of MALME-3 cells with BESPM resulted in an accumulation of N-acetylspermidine in cells and the enhanced excretion of putrescine, spermidine, and N-acetylspermidine into the medium. The relationship between SSAT induction and growth sensitivity was deduced to be a possible function of increased excretion of acetylated polyamines leading to enhanced polyamine pool depletion. The data suggest that, in cell types in which it occurs, unusually high increases in SSAT activity may serve as a determinant of growth sensitivity to bis-ethyl spermine analogues or, alternatively, as a target for appropriately designed chemotherapeutic strategies.

Acetyltransferases↗
PubMed1980

Polyamines and polyamine complexes: analytical comments.

Different methods of sample preparation for determination of di- and polyamines (putrescine, spermidine, spermine) and their conjugates in serum, plasma, and urine are discussed. Higher polyamine values obtained after hydrolysis of serum or plasma acidic extracts suggest the presence of polyamine conjugates. Additional polyamines might be bound to proteins and released by hydrolysis of the crude samples. Biological studies of bound polyamines in health and disease will require procedures for quantitative analysis. For the pertinent measurements, radioimmunoassays employing antibodies developed against isolated polyamine conjugates may prove useful.

Acetylation↗
PubMed1995

Antagonist properties of polyamines and bis(ethyl)polyamines at N-methyl-D-aspartate receptors.

The polyamine spermine has both stimulatory and inhibitory effects on N-methyl-D-aspartate (NMDA) receptors. At recombinant NMDA receptors, effects of spermine are dependent on the subunit composition of the receptor. In the present work we have used voltage-clamp recording to examine the effects of polyamines and bis(ethyl)polyamines on recombinant NMDA receptors expressed in Xenopus oocytes. The compounds that were studied include several bis(ethyl)polyamines that may be clinically useful as antitumor agents. A number of pentaamines and bis(ethyl)pentaamines were found to act as potent voltage-dependent antagonists at heteromeric NR1A/NR2A and NR1A/NR2B receptors, but not at NR1A/NR2C receptors. Antagonism was more pronounced in oocytes voltage-clamped at -80 mV than at -20 mV. Some polyamine analogs also potentiated responses to glutamate at NR1A/NR2B receptors at membrane potentials of -20 to +40 mV, but this effect required higher concentrations of polyamines than did inhibition seen at hyperpolarized membrane potentials. At NR1A/NR2A receptors the block seen with pentaamines and bis(ethyl)pentaamines, but not with spermine or bis(ethyl)spermine, was maximal at a membrane potential of -100 mV and was relieved at more negative as well as at more positive membrane potentials. This suggests that the mechanism of inhibition of NMDA receptors by pentaamines is different from that of spermine. Pentaamines may permeate the ion channel of NMDA receptors at very hyperpolarized membrane potentials and may be useful for studying the structural properties of NMDA receptor channels.

Animals↗
PubMed2003

Induction of phase 2 enzymes by serum oxidized polyamines through activation of Nrf2: effect of the polyamine metabolite acrolein.

The naturally occurring polycationic polyamines including putrescine, spermidine, and spermine play an important role in cell growth, differentiation, and gene expression. However, circulating polyamines are potential substrates for several oxidizing enzymes including copper-containing serum amine oxidase. These enzymes are capable of oxidizing serum polyamines to several toxic metabolites including aldehydes and H(2)O(2). In this study, we investigated the effects of polyamines as inducers of phase 2 enzymes and other genes that promote cell survival in a cell culture system in the presence of bovine serum. Spermidine and spermine (50 microM) increased NAD(P)H quinone oxidoreductase (NQO1) activity up to 3-fold in murine keratinocyte PE cells. Transcript levels for glutathione S-transferase (GST) A1, GST M1, NQO1, gamma-glutamylcysteine ligase regulatory subunit, and UDP-glucuronyltransferase 1A6 were significantly increased by spermidine and this effect was mediated through the antioxidant response element (ARE). The ARE from the mouse GST A1 promoter was activated about 9-fold by spermine and 5-fold by spermidine treatment, but could be inhibited by the amine oxidase inhibitor, aminoguanidine, suggesting that acrolein or hydrogen peroxide generated from polyamines by serum amine oxidase may be mediators for phase 2 enzyme induction. Elevations of ARE-luciferase expression and NQO1 enzyme activity by spermidine were not affected by catalase, while both were completely repressed by aldehyde dehydrogenase treatment. Direct addition of acrolein to PE cells induced multiple phase 2 genes and elevated nuclear levels of Nrf2, a transcription factor that binds to the ARE. Expression of mutant Nrf2 repressed the activation of the ARE-luciferase reporter by polyamines and acrolein. These results indicate that spermidine and spermine increase the expression of phase 2 genes in cells grown in culture through activation of the Nrf2-ARE pathway by generating the sulfhydryl reactive aldehyde, acrolein.

Acrolein↗
PubMed1992

alpha-Methyl polyamines: metabolically stable spermidine and spermine mimics capable of supporting growth in cells depleted of polyamines.

In order to assess the tolerance of the target enzyme spermine synthase for alpha-substituents on the aminopropyl moiety of the substrate spermidine, 1-methylspermidine (MeSpd, 2) was synthesized. It was determined that MeSpd is a poor substrate for spermine synthase and is not a substrate for spermidine N1-acetyltransferase, suggesting that alpha-methylated polyamines might be metabolically stable and therefore useful tools for studying polyamine effects in intact cells. On the basis of initial cellular results with 2, 1-methylspermine (MeSpm, 3) and 1,12-dimethylspermine (Me2Spm, 4) were also synthesized. When added to cells (L1210, SV-3T3, or HT29) depleted of both putrescine and spermidine by prior treatment with alpha-(difluoromethyl)ornithine (DFMO), these alpha-methylated polyamines were able to restore cell growth to that observed in the absence of DFMO. In accord with the enzyme data noted above, metabolic studies indicated a slow conversion of 2 to 3, but no metabolism of 4 in these cells. It was concluded from these results that the alpha-methylated polyamines are able to substitute for the natural polyamines spermidine and spermine in critical biochemical processes which involve polyamines for continued cell growth. In accord with the hypothesis, preliminary data indicate that MeSpd and Me2Spm are as effective as spermidine and spermine, respectively, in promoting the conversion of B-DNA to Z-DNA.

Animals↗
PubMed2001

Endogenous levels of polyamines in the organs of cucumber plant (Cucumis sativus) and factors affecting leaf polyamine contents.

Polyamine compositions of various organs from hydroponically cultivated cucumber plants (Cucumis sativus L. cv. Sharp-1) and factors affecting the leaf polyamine content were examined. Diamine putrescine was found most abundantly in the root, while a relatively large amount of spermine was detected in the reproductive organs such as the immature fruit and the calyx (+stamen). Spermidine was present at the highest level in rapidly growing tissues such as newly expanded leaf and fruit at an early developing stage, implying the possible involvement of spermidine in the growth and development of these young tissues. Polyamine content of cucumber leaves changed during the day. Especially, the putrescine content of upper leaves showed a striking decrease from the morning to the night. Alterations of leaf Ca or Mg content did not significantly affect leaf polyamine composition. On the other hand, abnormal cucumber leaves showed altered polyamine composition. Yellowing of the leaf intervein resulted in a striking decrease in spermidine content without a significant change in putrescine and spermine content. By contrast, the leaves infected with the phytopathogen, powdery mildew, showed decreased putrescine and increased spermine content in response to the degree of fungi infection. The possible usefulness of polyamines as a diagnostic marker of plant development and physiological disorder is discussed.

Journal Article↗
PubMed1988

Studies of non-metabolizable polyamines that support growth of SV-3T3 cells depleted of natural polyamines by exposure to alpha-difluoromethylornithine.

A number of synthetic polyamine derivatives that included five achiral gem-dimethylspermidines and two analogous tetramethylated spermines were tested for their abilities to serve as substrates for enzymes metabolizing polyamines and for their capacities to substitute for the natural polyamines in cell growth. It was found that none of the compounds were effective substrates for spermine synthase, and only one, namely 8,8-dimethylspermidine, was a substrate for spermidine/spermine N1-acetyltransferase. However, all of the spermidine derivatives and 1,1,12,12-tetramethylspermine were able to support the growth of SV-3T3 cells in which endogenous polyamine synthesis was prevented by the addition of alpha-difluoromethylornithine. These results suggest that either spermidine or spermine can support cell growth without the need for metabolic interconversion. In contrast with the result with 1,1,12,12-tetramethylspermine, 3,3,10,10-tetramethylspermine did not restore growth of polyamine-depleted SV-3T3 cells. Comparison of the properties of these derivatives may prove valuable in understanding the physiological role of polyamines.

Acetyltransferases↗
PubMed1982

Polyamines and root formation in mung bean hypocotyl cuttings : I. Effects of exogenous compounds and changes in endogenous polyamine content.

The effect of several polyamines (putrescine, spermidine, and spermine), their precursors (l-arginine and l-ornithine), and some analogs and metabolic inhibitors (l-canavanine, l-canaline, and methylglyoxal-bis [guanylhydrazone]) on root formation have been studied in mung bean (Vigna radiata [L.] Wilczek) hypocotyl cuttings.Exogenously applied polyamines did not promote adventitious root formation. Rooting was inhibited by l-canavanine and l-canaline, and this inhibition was reversed by the corresponding amino acids l-arginine and l-ornithine. Methylglyoxal-bis (guanylhydrazone), an inhibitor of S-adenosylmethionine decarboxylase and polyamine biosynthesis, was also found to inhibit root formation. All compounds at concentrations of >10(-4) molarity completely inhibited natural root formation, whereas at <10(-5) molarity only the indole-butyric acid-induced root formation was inhibited.Indole-butyric acid-induced root formation was accompanied by a considerable increase in polyamine levels, more than 2-fold of the control. Whereas senescing (unrooted) cuttings evinced a rapid decline in polyamine content during 48 hours, indole-butyric acid treatment resulted in elevated levels of putrescine and increased putrescine to spermidine ratio. The changes in polyamines were dependent on indole-butyric acid concentration and were organ specific.

Journal Article↗
PubMed1988

Endogenous polyamines associate with DNA during its condensation in mammalian tissue. A fluorescence cytochemical and immunocytochemical study of polyamines in fetal rat liver.

The polyamines spermidine and spermine are essential for cell proliferation and differentiation. By two independent fluorescence cytochemical methods as well as by immunocytochemistry, we have studied the distribution of these molecules in fetal rat liver. Strong reactions for polyamines were found in highly condensed chromatin, present in chromosomes in mitotic cells, and in condensed nuclei in late erythropoietic cells. Moreover, polyamines were so closely associated with DNA in condensed chromatin that DNase pretreatment was necessary for making them available for reaction with antibodies. In other cells, polyamines were mainly localized to the cytoplasm. Studies of cells at different stages in erythropoiesis revealed that polyamines become associated with DNA during its condensation and inactivation. Our data strongly indicate that polyamines participate in the condensation of DNA.

Animals↗
PubMed1993

Effect of a bis(benzyl)polyamine analogue, and DL-alpha-difluoromethylornithine on parasite suppression and cellular polyamine levels in golden hamster during Leishmania donovani infection.

We examined the antileishmanial activity of DL-alpha-difluoromethylornithine (DFMO) and a bis(benzyl)polyamine analogue (MDL 27695; N,N'-bis (3-[(phenyl-methyl)amino] propyl) 1,7-diaminoheptane) in L. donovani infected golden hamsters. DFMO, an enzyme activated irreversible inhibitor of ornithine decarboxylase, the rate limiting enzyme in polyamine biosynthesis, has potent antileishmanial activity. When given as a 2% solution in drinking water 2 days after infection and continued for 4 days, it suppressed liver parasites by 90% and spleen parasites by 99%. Liver parasites were suppressed by 50% and spleen parasites by 77% in L. donovani infected hamsters when treated three times per day for 4 days with a total dose of 60 mg kg-1 body weight of MDL 27695. The polyamine content of the liver and spleen of hamsters was determined after 8 days of L. donovani infection and also after treatment with these drugs. Putrescine and spermidine levels increased significantly in both liver and spleen after Leishmania infection of golden hamsters. Treatment with drugs that inhibit the growth of Leishmania mastigotes in the liver and spleen of golden hamsters also reduced polyamine levels of previously infected golden hamsters. There is a close correlation between the therapeutic activity of the drugs and the polyamine content.

Animals↗
PubMed1999

Effects of MDL 72527, a specific inhibitor of polyamine oxidase, on brain edema, ischemic injury volume, and tissue polyamine levels in rats after temporary middle cerebral artery occlusion.

The possible effects of the polyamine interconversion pathway on tissue polyamine levels, brain edema formation, and ischemic injury volume were studied by using a selective irreversible inhibitor, MDL 72527, of the interconversion pathway enzyme, polyamine oxidase. In an intraluminal suture occlusion model of middle cerebral artery in spontaneously hypertensive rats, 100 mg/kg MDL 72527 changed the brain edema formation from 85.7 +/- 0.3 to 84.5 +/- 0.9% in cortex (p < 0.05) and from 79.9 +/- 1.7 to 78.4 +/- 2.0% in subcortex (difference not significant). Ischemic injury volume was reduced by 22% in the cortex (p < 0.05) and 17% in the subcortex (p < 0.05) after inhibition of polyamine oxidase by MDL 72527. There was an increase in tissue putrescine levels together with a decrease in spermine and spermidine levels at the ischemic site compared with the nonischemic site after ischemia-reperfusion injury. The increase in putrescine levels at the ischemic cortical and subcortical region was reduced by a mean of 45% with MDL 72527 treatment. These results suggest that the polyamine interconversion pathway has an important role in the postischemic increase in putrescine levels and that blocking of this pathway can be neuroprotective against neuronal cell damage after temporary focal cerebral ischemia.

Animals↗
PubMed2001

Cloning and characterization of a human polyamine oxidase that is inducible by polyamine analogue exposure.

Mammalian polyamine catabolism is under the control of two enzymes, spermidine/spermine N1-acetyltransferase and the flavin adenine dinucleotide-dependent polyamine oxidase (PAO). In this study, the cloning and initial characterization of human PAO is reported. A 1894-bp cDNA with an open reading frame of 1668-bp codes for a protein of 555 amino acids. In vitro transcription/translation of this cDNA clone produces the expected M(r) 61,900 protein with PAO activity. The PAO activity of this clone is inhibited by MDL 72,527, a specific inhibitor of mammalian PAO. However, neither pargyline, a specific monoamine oxidase inhibitor, nor semicarbazide, a specific diamine oxidase inhibitor, inhibits the PAO activity of this clone. PAO has been referred to as being constitutively expressed. However, 24-h exposure of a non-small cell lung carcinoma cell line, NCI H157, to 10 microM of N1,N"-bis(ethyl)norspermine results in approximately 5-fold induction of PAO mRNA and a >3-fold induction of PAO activity. These results demonstrate that in at least one cell type, PAO is up-regulated in response to polyamine analogue exposure. The PAO clone described here should provide a useful tool, which will facilitate the dissection of the role of polyamine catabolism in normal growth and in response to the antitumor polyamine analogues.

Amino Acid Sequence↗
PubMed2002

Detoxification of the polyamine analogue N1-ethyl-N11-[(cycloheptyl)methy]-4,8-diazaundecane (CHENSpm) by polyamine oxidase.

PURPOSE: Analogues of the naturally occurring polyamines, alkylated on both terminal amines, are being developed as anticancer drugs. Because bisalkylated derivatives of putrescine (1,4-diaminobutane) are potent inhibitors of the flavin adenine dinucleotide-dependent polyamine oxidase (PAO), we asked whether PAO could detoxify synthetic bisalkylated polyamines with chain lengths longer than putrescine. EXPERIMENTAL DESIGN: We investigated the effects of one polyamine analogue in Chinese hamster ovary (CHO) and HCT116 human colon tumor-derived cells, which express dramatically different levels of PAO activity, and in these same cells treated with an inhibitor of PAO. RESULTS: Concentrations of N1-ethyl-N11-[(cycloheptyl)methyl]-4,8-diazaundecane (CHENSpm), ranging from 0.3 to 10 microM, caused growth arrest and reduced cell survival in HCT116 cells but not in CHO cells. This cell line-specific difference in CHENSpm toxicity was not attributable to differences in analogue uptake, because intracellular levels of the drug were similar in CHO and HCT116 cells treated with equivalent concentrations of CHENSpm in the presence of MDL 72,527, a specific inhibitor of PAO. The PAO inhibitor, in combination with CHENSpm, caused a significant increase in intracellular CHENSpm levels and increased growth inhibition and cell damage in CHO cells but not in HCT116 cells. CHO cells, but not HCT116 cells, produced two primary amine-containing metabolites from CHENSpm that were suppressed by MDL 72,527. CONCLUSIONS: These data demonstrate that CHENSpm is detoxified in PAO-expressing CHO cells, but not in PAO-deficient HCT116 cells, by a mechanism yielding products containing free primary amine groups and implicate PAO as the detoxification enzyme. Because other studies suggest that PAO may be self-regulated in some tumors, differential expression of PAO may be the basis for selective toxicity of CHENSpm and other N-substituted polyamine analogues in certain cancers.

Animals↗
PubMed1989

Urinary polyamine levels in cancer patients treated with D,L-alpha-difluoromethylornithine, an inhibitor of polyamine biosynthesis.

The polyamine biosynthesis inhibitor D,L-alpha-Difluoromethylornithine hydrochloride monohydrate (DFMO) has cytostatic and cytotoxic effects against various human tumor cell lines in vitro. We measured levels of the polyamines putrescine and spermidine in the urine of cancer patients undergoing "conventional" chemotherapy in a two-arm randomized phase I-II study with and without additional DFMO administered orally at a dose of 1.7 g/sq.m. t.i.d. The study group included 38 patients with carcinoma of the breast, stomach, prostate, or female genital organs or metastatic carcinoma of unknown origin. A control group of 32 patients with similar malignancies received "conventional" chemotherapy without DFMO. Polyamine levels were determined periodically in the urine of all patients. In DFMO-treated patients, a significant decrease in putrescine and spermidine levels was observed after 3 weeks of DFMO therapy (the first time point evaluated) that usually persisted throughout the course of treatment. Significant differences in polyamine levels between DFMO-treated and control patients were observed for patients in remission. Less significant differences were noted, however, for patients with static or progressive disease between DFMO-treated and control groups. DFMO activity appears to be reflected by a long-term decrease in urinary polyamine levels.

Antineoplastic Combined Chemotherapy Protocols↗
PubMed1985

Regulation of polyamine biosynthesis by antizyme and some recent developments relating the induction of polyamine biosynthesis to cell growth. Review.

This review considers the role of antizyme, of amino acids and of protein synthesis in the regulation of polyamine biosynthesis. The ornithine decarboxylase of eukaryotic cells and of Escherichia coli can be non-competitively inhibited by proteins, termed antizymes, which are induced by di- and poly- amines. Some antizymes have been purified to homogeneity and have been shown to be structurally unique to the cell of origin. Yet, the E. coli antizyme and the rat liver antizyme cross react and inhibit each other's biosynthetic decarboxylases. These results indicate that aspects of the control of polyamine biosynthesis have been highly conserved throughout evolution. Evidence for the physiological role of the antizyme in mammalian cells rests upon its identification in normal uninduced cells, upon the inverse relationship that exists between antizyme and ornithine decarboxylase as well as upon the existence of the complex of ornithine decarboxylase and antizyme in vivo. Furthermore, the antizyme has been shown to be highly specific; its Keq for ornithine decarboxylase is 1.4 X 10(11) M-1. In addition, mammalian cells contain an anti-antizyme, a protein that specifically binds to the antizyme of an ornithine decarboxylase-antizyme complex and liberates free ornithine decarboxylase from the complex. In E. coli, in which polyamine biosynthesis is mediated both by ornithine decarboxylase and by arginine decarboxylase, three proteins (one acidic and two basic) have been purified, each of which inhibits both these enzymes. They do not inhibit the biodegradative ornithine and arginine decarboxylases nor lysine decarboxylase. The two basic inhibitors have been shown to correspond to the ribosomal proteins S20/L26 and L34, respectively. The relationship of the acidic antizyme to other known E. coli proteins remains to be determined. In mammalian cells, ornithine decarboxylase can be induced by a broad spectrum of compounds. These range from hormones and growth factors to natural amino acids such as asparagine and to non-metabolizable amino acid analogues such as alpha-amino-isobutyric acid. The amino acids that induce ornithine decarboxylase as well as those that promote polyamine uptake utilize the sodium dependent A and N transport systems. Consequently, they act in concert and increase intracellular polyamine levels by both mechanisms. The induction of ornithine decarboxylase by growth factors, such as NGF, EGF, and PDGF as well as by insulin requires the presence of these same amino acids and does not occur in their absence. However, the inducing amino acid need not be incorporated into protein nor covalently modified.(ABSTRACT TRUNCATED AT 400 WORDS)

Amino Acids↗
PubMed2004

A molecular dynamics simulation study of polyamine- and sodium-DNA. Interplay between polyamine binding and DNA structure.

Four different molecular dynamics (MD) simulations have been performed for infinitely long ordered DNA molecules with different counterions, namely the two natural polyamines spermidine(3+) (Spd3+) and putrescine(2+) (Put2+), the synthetic polyamine diaminopropane(2+) (DAP2+), and the simple monovalent cation Na+. All systems comprised a periodical hexagonal cell with three identical DNA decamers, 15 water molecules per nucleotide, and counterions balancing the DNA charge. The simulation setup mimics the DNA state in oriented DNA fibers, previously studied using NMR and other experimental methods. In this paper the interplay between polyamine binding and local DNA structure is analyzed by investigating how and if the minor groove width of DNA depends on the presence and dynamics of the counterions. The results of the MD simulations reveal principal differences in the polyamine-DNA interactions between the natural [spermine(4+), Spd3+, Put2+] and the synthetic (DAP2+) polyamines.

Base Sequence↗
PubMed1986

Inhibition of the respiratory burst of human neutrophils by the polyamine oxidase-polyamine system.

The addition of the polyamines, spermine and spermidine, to human neutrophils caused a depression of the hexose-monophosphate (HMP) shunt activity of neutrophils stimulated with latex particles but not of unstimulated cells. The effect was dependent on the presence of bovine serum and was not observed when normal human serum was substituted for bovine serum. The polyamine oxidase (PAO) in bovine serum was probably responsible for generating the activity since normal human serum lacks PAO. A role for PAO was further supported by the finding that partially purified bovine PAO in the presence of polyamines similarly mediated inhibition of HMP shunt activity in stimulated neutrophils. Catalase failed to prevent the inhibitory effects of the PAO-polyamine system suggesting that H2O2 is not the responsible product. In addition, our results show that human pregnancy serum known to contain PAO activity in the presence of polyamines mediated a similar inhibition of the respiratory burst.

Animals↗
PubMed1986

Solid-phase extraction and determination of dansyl derivatives of unconjugated and acetylated polyamines by reversed-phase liquid chromatography: improved separation systems for polyamines in cerebrospinal fluid, urine and tissue.

A sensitive and simple liquid chromatographic assay with fluorometric detection for unconjugated and acetylated polyamines in biological fluids is described. After precolumn derivatization with dansyl chloride, unconjugated polyamines and acetylated polyamines were extracted by elution from a Bond-Elut C18 column and then separated on a reversed-phase column with gradient elution. The complete analysis of unconjugated putrescine, spermidine, and spermine in either hydrolyzed urine, cerebrospinal fluid or tissue could be accomplished within 20-26 min, while the simultaneous analysis of unconjugated polyamines and monoacetylpolyamines could be completed within 40 min. Unhydrolyzed urine and cerebrospinal fluid required a Bond-Elut cation-exchange clean-up before dansylation. Standard curves for the assay were linear up to 20 nmol/ml, and the within-day and day-to-day coefficients of variation were between 1.1 and 4.6% and between 1.6 and 11.8%, respectively. Results obtained with the method were compared with results obtained with a well established modified amino acid analyzer method for urine, tissue and cerebrospinal fluid samples. The correlation coefficients between these two methods were in the range 0.933-0.996. Detection limits between 50 and 150 fmol were achieved for unconjugated and acetylated polyamines. Of more than twenty drugs and amines tested for possible interference with the assay, only normetanephrine was found to have the same retention time as the internal standard 1,6-diaminohexane.

Acetylation↗