NaCl-Induced Changes in Putrescine Content and Diamine Oxidase Activity in Roots of Rice Seedlings

The effects of NaCl on putrescine (Put) content and diamine oxidase (DAO) activity in roots of rice seedlings were examined. NaCl treatment lowered the content of Put and increased the activity of DAO in roots. Our current results indicate that Cl^– is not required for NaCl-induced decline in Put content and increase in DAO activity in roots. Put content in roots of rice seedlings exposed to NaCl is possibly regulated by DAO activity.     

Quantification of gut injury with diamine oxidase activity: development of a fission neutron RBE and measurements with combined injury in mouse models

Plasma and small intestine diamine oxidase (DAO) activities were measured on Days 2, 4, and 6 following irradiation of mice with a range of doses of fission neutrons and 60Co. With increasing doses of radiation, plasma DAO activity increased on Day 2 and intestinal DAO activity decreased on Day 4; moreover, the approximate relative biological effectiveness values for these changes in activity were 5.81 for plasma DAO activity on Day 2 and 3.88 for intestinal DAO activity on Day 4. On Day 6 relatively high levels of radiation caused DAO activity in the small intestine to remain depressed whereas low levels resulted in recovery with activities at or near controls. In animals with combined injury (radiation plus 30% surface burn or wound), changes in DAO activity in the intestine were similar to those with radiation alone; plasma DAO activity, in contrast to radiation alone, did not show an increase at the 2-day mark. These dose-dependent relationships should provide a basis for using DAO as a potential indicator of biological damage from radiation exposure within the lethal range.     

Developmental aspects of polyamine-oxidizing enzyme activities in the mouse kidney. Effects of testosterone

Summary In the present study developmental patterns of renal polyamineoxidizing enzymes polyamine oxidase (PAO) and diamine oxidase (DAO) in male and female ICR mice were demonstrated. The effects of testosterone (10g/100g body weight) on renal PAO and DAO activities were also studied. The differences between sexes in both PAO and DAO activities were most clearly expressed in the immature kidney. At the age of 20 days PAO and DAO activities were 1.52 fold (p < 0.01) and 1.75 (p < 0.02) respectively higher in male mouse kidney than in female. Maturational processes reflected in significant increases in polyamine- oxidizing enzyme activities mainly in female mouse kidney, comparable with the gain in the kidney wet weight. Our data show that testosterone is able to influence renal PAO and DAO activities in addition to the well-known stimulation of polyamine biosynthesis. The hormonal effects were sex and age dependent. The influence of testosterone on renal PAO activity was mainly age dependent. The slight stimulation of renal PAO activity observed in 20- and 50-day old mice, 24h after testosterone administration, change with a decrease in the enzyme activity at the age of 70 days. The effects of testosterone on renal DAO activity were mainly sex dependent. Testosterone caused stimulation of DAO activity with a very close magnitude (nearly twice) in female mouse kidney, independently of the age of mice. In contrast, in male mice the hormone treatment resulted in a statistically significant increase in renal DAO activity at the age of 70 days (.1.3 fold, p < 0.05) only. It could be suggested that our data indicate the different contribution of renal PAO and DAO in androgen regulation of polyamine levels, depending on sex and the stage of the postnatal development.     

Molecular characterization of D-amino acid oxidase from common carp Cyprinus carpio and its induction with exogenous free D-alanine

A full-length cDNA encoding D-amino acid oxidase (DAO, EC 1.4.3.3) was cloned and sequenced from the hepatopancreas of carp fed a diet supplemented with D-alanine. This clone contained an open reading frame encoding 347 amino acid residues. The deduced amino acid sequence exhibited about 60 and 19-29% identity to mammalian and microbial DAOs, respectively. The expression of full-length carp DAO cDNA in Escherichia coli resulted in a significant level of protein with DAO activity. In carp fed the diet with D-alanine for 14 days, DAO mRNA was strongly expressed in intestine followed by hepatopancreas and kidney, but not in muscle. During D-alanine administration, DAO gene was expressed quickly in hepatopancreas with the increase of DAO activity. The inducible nature of carp DAO indicates that it plays an important physiological role in metabolizing exogenous D-alanine that is abundant in their prey invertebrates, crustaceans, and mollusks.     

Higher accumulation of gamma-aminobutyric acid induced by salt stress through stimulating the activity of diamine oxidases in Glycine max (L.) Merr. roots.

Polyamines (PAs) are assumed to perform their functions through their oxidative product such as gamma-aminobutyric acid (GABA) formation. However, there is only limited information on the interrelation between PA degradation and GABA accumulation under salt stress. In order to reveal a quantitative correlation between PA oxidation and GABA accumulation, the effects of treatments with different NaCl concentrations, along with aminoguanidine (AG, a specific inhibitor of diamine oxidases (DAO; EC: 1.4.3.6)) and a recovery test from salt stress on endogenous free PAs, gamma-aminobutyric acid (GABA) accumulation and DAO activity were determined in roots of soybean [Glycine max (L.) Merr.] cultivar Suxie-1. The results showed that the levels of putrescine (Put), cadaverine (Cad), and spermidine (Spd) decreased significantly with increasing salt concentrations. This occurred because salt stress strongly promoted DAO activity to stimulate PA degradation. GABA accumulation increased with growing NaCl concentrations, about an 11- to 17-fold increase as compared with the control plants. AG treatment increased the accumulation of endogenous free PAs as a result of a strong retardation of DAO activity, but decreased GABA accumulation. The recovery for 6 days in 1/2 Hoagland solution from 100mM NaCl stress resulted in a decrease in DAO activity, a rebound of PA levels and a simultaneous reduction of GABA content. A close correlation was observed between the changes in DAO activity and GABA accumulation. The results indicated that higher GABA accumulation (about 39%) induced by salt stress could come from PA degradation, suggesting that PAs might perform their functions through GABA formation under salt stress.     

Voluntary exercise stabilizes photic entrainment of djungarian hamsters (Phodopus sungorus) with a delayed activity onset

ABSTRACT

The Djungarian hamsters of our breeding colony show unstable daily activity patterns when kept under standard laboratory conditions. Moreover, part of them develops a delayed activity onset (DAO) or an arrhythmic phenotype. In former studies, we have shown that the system of photic entrainment works at its limits. If the period length (tau) increases, which is the case in DAO hamsters, the light-induced phase advances are too small to compensate the daily delay of the activity rhythm caused by tau being longer than 24 h. Accordingly, under natural conditions, there must be further (environmental) factors to enable a stable entrainment. One of these may be the higher level of motor activity. Animals must cover long distances to search for food, sexual partners and others. In the laboratory, hamsters are kept singly in small cages. This does restrict animals’ options for motor activity. Also, there is less need for moving around as the hamsters are fed ad libitum.

In the present study, a series of experiments was performed to investigate the putative effect of the activity level. To begin with, wild type (WT) and DAO animals were given access to running wheels. 50% of DAO hamsters developed a WT activity pattern. As the main reason for the DAO phenomenon is their long tau together with a too weak photic phase response, the effect of wheel running on these parameters was investigated in further experiments. With higher activity level, tau decreased in WT hamsters but increased in DAO animals even though the increase for the activity onset was only close to significance. Moreover, the photic phase responses were weaker though significant only for the activity offset of DAO hamsters.

Based on the assumptions that running wheel activity will affect the phase response and/or the free running period, the results of the present paper do not provide an explanation for why part of DAO hamsters developed a WT phenotype when they had access to running wheels. Obviously, mechanisms downstream from the suprachiasmatic nuclei must be taken into account when investigating the stabilizing, improving circadian entrainment effect of motor activity.     

γ-氨基丁酸对低氧胁迫下甜瓜幼苗多胺代谢的影响

以‘西域一号’甜瓜为试验材料,采用营养液水培法,研究了低氧胁迫下外源添加γ-氨基丁酸(GABA)对甜瓜幼苗多胺代谢的影响.结果表明:与通气对照相比,低氧胁迫处理的甜瓜幼苗谷氨酸脱羧酶(GAD)活性和GABA含量显著提高,同时多胺合成酶活性提高诱导多胺含量显著增加,但二胺氧化酶(DAO)和多胺氧化酶(PAO)活性也显著提高;根系精氨酸脱羧酶(ADC)活性提高幅度较大,导致根系游离态腐胺含量较高,而叶片乌氨酸脱羧酶(ODC)和S-腺苷甲硫氨酸脱羧酶(SAMDC)活性提高幅度较大,导致叶片游离态亚精胺(Spd)含量较高;根系游离态DAO和PAO活性显著低于叶片,其细胞壁结合态PAO活性显著高于叶片.与低氧胁迫处理相比,低氧胁迫下外源添加GABA处理的甜瓜幼苗叶片和根系中GABA和谷氨酸含量均显著提高,而GAD活性显著降低;精氨酸、鸟氨酸、甲硫氨酸含量的提高促使多胺合成酶活性显著提高,从而诱导多胺含量显著增加,DAO和PAO活性显著降低.     

Distribution of polyamines and their related catabolic enzyme in etiolated and light-grown leguminosae seedlings

Diamine-oxidase (DAO; EC 1.4.3.6) activity and di-and polyamine levels were estimated along the epicotyl and root of light-grown and etiolated lentil (Lens culinaris Medicus) and pea (Pisum sativum L.) seedlings. The activity of DAO was higher in etiolated epicotyls than in lightgrown ones. In both species there was a positive correlation between DAO activity and the diamine (putrescine and cadaverine) levels along the whole epicotyl and root. Polyamine (spermine and spermidine) distribution seemed to be associated with the meristematic and elongating zone of the epicotyl and root. The physiological function of DAO is discussed in relation to its possible role in providing hydrogen peroxide to peroxidase-dependent reactions occurring in the cell wall.Abbreviations CAD cadaverine - DA diamine - DAO diamine oxidase - PA polyamine - PUT putrescine - SPD spermidine - SPM spermine     

Radiation effects on diamine oxidase activities in intestine and plasma of the rat

Diamine oxidase (DAO; EC 1.4.3.6) activity was measured in plasma and in ileal tissue homogenates prepared from male Sprague-Dawley rats euthanized at 1-15 days after acute whole-body irradiation with 14.5-MeV electrons. Animals irradiated with 1 Gy showed no diminution in plasma and ileal DAO activities through Day 13 relative to nonirradiated controls. Animals irradiated with 5, 10, and 12 Gy displayed marked declines in ileal DAO activity, with levels reaching a nadir on Day 3. This was paralleled by a decrease in plasma DAO activity in all three dose groups. Recovery of ileal and plasma DAO levels was later seen as early as Day 4 in animals irradiated with 5- and 10-Gy doses, but animals receiving 12 Gy did not survive beyond Day 3. The relationship between radiation dose and levels of plasma and ileal DAO on Day 3, the time of maximum decrease at all doses, was also investigated. Ileal DAO activity decreased almost linearly between 2 and 8 Gy. Plasma DAO activity closely paralleled the dose dependency of the ileal levels. These data suggest that plasma DAO activity might be useful as a biologic marker of intestinal epithelial injury and recovery after acute radiation exposure.     

Osmotic stress-induced changes in cell wall peroxidase activity and hydrogen peroxide level in roots of rice seedlings

The changes in activity of peroxidase (POD) extracted from the cellwalls and the level of H₂O₂ in rice seedling rootstreatedwith mannitol and their correlation with root growth were investigated.Increasing concentrations of mannitol from 92 to 276 mM, which isiso-osmotic with 50 to 150 mM NaCl, progressively reduced rootgrowth and increased POD activities extracted from the cell walls of riceroots.The reduction of growth was also correlated with an increase inH₂O₂ level. Both diamine oxidase (DAO) and NADHperoxidase(NADH-POD) are known to be responsible for the generation ofH₂O₂. Mannitol treatment increased DAO but not NADH-PODactivities in roots of rice seedlings, suggesting that DAO contributes to thegeneration of H₂O₂ in the cell walls of mannitol-treatedroots. An increase in the level of H₂O₂ and the activityof POD extracted from the cell walls of rice roots preceded root growthreduction caused by mannitol. An increase in DAO activity coincided with anincrease in H₂O₂ in roots caused by mannitol. Since DAOcatalyses the oxidation of putrescine, the demonstration that mannitolincreasesthe activity of DAO in roots is consistent with those that mannitol decreasesthe level of putrescine. In conclusion, cell-wall stiffening catalysed by PODispossibly involved in the regulation of root growth reduction caused bymannitol.     

Low serum diamine oxidase (DAO) activity levels in patients with migraine

Histamine intolerance is a disorder in the homeostasis of histamine due to a reduced intestinal degradation of this amine, mainly caused by a deficiency in the enzyme diamine oxidase (DAO). Among the several multi-faced symptoms associated with histamine intolerance, headache is one of the most recognized and disabling consequences. The aim of this study was to determine the prevalence of DAO deficiency in patients with a confirmed migraine diagnosis according to the current International Headache Society (IHS) and in non-migraine subjects. DAO activity was assessed in a total of 198 volunteers recruited at the Headache Unit of the Hospital General de Catalunya, 137 in the migraine group and 61 as a control group. DAO enzyme activity in blood samples was determined by ELISA test. Values below 80 HDU/ml (Histamine Degrading Unit/ml) were considered as DAO deficient. Mean value of DAO activity from migraine population (64.5 ± 33.5 HDU/ml) was significantly lower (p < 0.0001) than that obtained from healthy volunteers (91.9 ± 44.3 HDU/ml). DAO deficiency was more prevalent in migraine patients than in the control group. A high incidence rate of DAO deficiency (87%) was observed in the group of patients with migraine. On the other hand, 44% of non-migranous subjects had levels of DAO activity lower than 80 HDU/ml. Despite the multifactorial aetiology of migraine, these results seem to indicate that this enzymatic deficit could be related to the onset of migraine.     

Potential role for astroglial D-amino acid oxidase in extracellular D-serine metabolism and cytotoxicity

D-amino acid oxidase (DAO) is a flavoenzyme that catalyzes the oxidation of D-amino acids. In the brain, gene expression of DAO is detected in astrocytes. Among the possible substrates of DAO in vivo, D-serine is proposed to be a neuromodulator of the N-methyl-D-aspartate (NMDA) receptor. In a search for the physiological role of DAO in the brain, we investigated the metabolism of extracellular D-serine in glial cells. Here we show that after D-serine treatment, rat primary type-1 astrocytes exhibited increased cell death. In order to enhance the enzyme activity of DAO in cells, we established stable rat C6 glial cells overexpressing mouse DAO designated as C6/DAO. Treatment with a high dose of D-serine led to the production of hydrogen peroxide (H(2)O(2)) followed by apoptosis in C6/DAO cells. Among the amino acids tested, D-serine specifically exhibited a significant cell death-inducing effect. DAO inhibitors, i.e., sodium benzoate and chlorpromazine, partially prevented the death of C6/DAO cells treated with D-serine, indicating the involvement of DAO activity in d-serine metabolism. Overall, we consider that extracellular D-serine can gain access to intracellular DAO, being metabolized to produce H(2)O(2). These results support the proposal that astroglial DAO plays an important role in metabolizing a neuromodulator, D-serine.     

A single-base-pair substitution abolishes D-amino-acid oxidase activity in the mouse.

Mutant ddY/DAO- mice lacking D-amino-acid oxidase (DAO) activity were examined for the cause of their lack of enzyme activity. Total RNA was extracted from the kidney of the ddY/DAO- mice and cDNA was synthesized. After cDNA encoding DAO was amplified by the polymerase chain reaction it was cloned into a plasmid and sequenced. Comparison of the DAO cDNA sequence with that of normal BALB/c mice revealed the presence of a single-base substitution (G----A) which causes a Gly-181----Arg substitution in the middle of the enzyme molecule. The mutant DAO cDNA was inserted into an expression vector and was expressed in transfected COS-1 cells. The transfected cells synthesized the DAO mutant protein, but they did not show DAO activity. In contrast, when cells were transfected with an expression vector carrying wild-type DAO cDNA, where the substituted base-pair was replaced by a normal base-pair, they showed DAO activity. These results indicate that the single base-pair substitution is the cause of the loss of DAO activity in the ddY/DAO- mice.     

γ-氨基丁酸对低氧胁迫下甜瓜幼苗多胺代谢的影响

以‘西域一号’甜瓜为试验材料,采用营养液水培法,研究了低氧胁迫下外源添加γ-氨基丁酸(GABA)对甜瓜幼苗多胺代谢的影响.结果表明：与通气对照相比,低氧胁迫处理的甜瓜幼苗谷氨酸脱羧酶(GAD)活性和GABA含量显著提高,同时多胺合成酶活性提高诱导多胺含量显著增加,但二胺氧化酶(DAO)和多胺氧化酶(PAO)活性也显著提高;根系精氨酸脱羧酶(ADC)活性提高幅度较大,导致根系游离态腐胺含量较高,而叶片鸟氨酸脱羧酶(ODC)和S-腺苷甲硫氨酸脱羧酶(SAMDC)活性提高幅度较大,导致叶片游离态亚精胺(Spd)含量较高;根系游离态DAO和PAO活性显著低于叶片,其细胞壁结合态PAO活性显著高于叶片.与低氧胁迫处理相比,低氧胁迫下外源添加GABA处理的甜瓜幼苗叶片和根系中GABA和谷氨酸含量均显著提高,而GAD活性显著降低;精氨酸、鸟氨酸、甲硫氨酸含量的提高促使多胺合成酶活性显著提高,从而诱导多胺含量显著增加,DAO和PAO活性显著降低.     

Effects of prostaglandins on peripheral progesterone and uterine diamine oxidase in the pregnant hamster

Serum progesterone and uterine levels of diamine oxidase (DAO) activity were determined during pregnancy in hamsters. Progesterone was elevated on Day 1 of pregnancy, had a transient peak on Day 5, remained relatively constant on Days 6–10, and then increased on Days 13 and 14. Uterine DAO activity could not be detected until Day 7 of pregnancy, approximately 1 $\text{1}\text{2}$ days after the initiation of implantation. DAO activity was associated with placental tissue, and more than 90% of the activity was localized in the maternal placenta. The temporal relationship between changes in serum concentrations of progesterone and uterine levels of DAO activity following PG administration also was studied. Serum progesterone was significantly depressed by 6 hr after treatment with PGs on Day 7 of pregnancy. However, uterine levels of DAO activity at 6 hr in the treated animals were not different from those in control animals. In contrast, both the serum progesterone concentrations and uterine levels of DAO activity were significantly lower at 24 hr after PG treatment. The effects of PG treatment on uterine DAO activity were completely blocked by concomitant administration of progesterone. However, concomitant administration of Provera® only blocked the effect of one PG analog that was tested (9-deoxo-9-methylene-16,16-dimethyl0-PGE₂). The data indicate that changes in uterine DAO activity following treatment with the PGs used here are primarily a consequence of a decrease in peripheral progesterone (i.e. a luteolytic effect of the PG).     

Association of diamine oxidase and <Emphasis Type="Italic">S</Emphasis>-adenosylhomocysteine hydrolase in <Emphasis Type="Italic">Nicotiana tabacum</Emphasis> extracts

The oxidative deamination of methylated putrescine by a diamine oxidase activity (DAO) is an important step in the biosynthesis of nicotine in tobacco and tropane alkaloids in several Solanaceous plants. A polyclonal rabbit antiserum was previously developed to a purported purified DAO enzyme from Nicotiana tabacum. The antiserum bound to a single 53 kDa protein and immunoprecipitated 80 of DAO activity from tobacco root extracts. In an effort to obtain DAO cDNAs, this antiserum was used to screen a tobacco cDNA expression library and three distinct immunoreactive cDNA clones were isolated. These cDNAs encoded predicted proteins that were either identical or nearly identical to predicted S-adenosylhomocysteine hydrolase (SAHH) from two Nicotiana species. Thus, the rabbit antiserum was not specific to DAO, even though it immunodepleted the majority of DAO activity from root extracts. Alternative hypotheses to explain the DAO immunodepletion results (such as poisoning of DAO activity or that SAHH is a bifunctional enzyme) were tested and ruled out. Therefore, we hypothesize that SAHH associates with DAO as part of a larger multienzyme complex that may function in planta as a nicotine metabolic channel.     

Production of D-amino acid oxidase (DAO) of Trigonopsis variabilis in Schizosaccharomyces pombe and the characterization of biocatalysts prepared with recombinant cells

The cDNA of D-amino acid oxidase (DAO) gene isolated from Trigonopsis variabilis was expressed in Schizosaccharomyces pombe. A clone, ASP327-10, transformed with plasmid vector, pTL2M5DAO, expressed catalytically active DAO in the presence of G418, and converted Cephalosprin C to alpha-ketoadipyl-7-cephalosporanic acid (KA-7-ACA) and glutaryl-7-aminocephalosporanic acid (GL-7-ACA). Biocatalysts were prepared using ASP327-10 and T. variabilis, and evaluated to demonstrate the feasibility of recombinant S. pombe for industrial application. The cells were immobilized by crosslinking polyethylene imine after glutardialdehyde (GDA) fixation and permeabilization by alkaline treatment. Although the biocatalyst prepared from ASP327-10 exhibited DAO activity, catalase activity still remained fully even after permeabilization, under which condition, the catalase activity of T. variabilis decreased to 20-30%. Heat treatment was required before cell fixation by GDA to inactivate the catalase in S. pombe. This improved the efficiency of bioconversion to GL-7-ACA, but caused poor mechanical strength in the biocatalyst of S. pombe. To overcome this weakness, a catalase-deficient host strain was obtained by ethylmethansulfate mutagenesis. Moreover, taking economics into consideration, the integrative vector, pTL2M5DAO-8XL, with multi-copies of expression cassette was constructed to express DAO in S. pombe even in the absence of G418. The newly established integrant, ASP417-7, did not exhibit any catalase activity so that heat treatment was not required. The obtained integrant and its biocatalyst were significantly improved in GL-7ACA conversion ability and mechanical strength. This study demonstrates that the established integrant is a potential candidate as an alternative source of DAO enzyme.     

Diamine oxidase is involved in H(2)O(2) production in the chalazal cells during barley grain filling

The localization and activities of diamine oxidase (DAO, EC 1.4.3.6) and polyamine oxidase (PAO, EC 1.4.3.4) together with polyamine levels have been investigated in developing grains of barley (Hordeum vulgare L.). DAO (pH 7.5) is present mainly in vascular tissue and its neighbouring cells, namely chalazal cells and nucellar projection, while PAO (pH 6.0) is mainly localized in the chlorenchymatous cells of the crease and at the base of the vascular tissue. Activities of both these enzymes appear to be independently-regulated, as DAO activity increased steadily throughout grain development while PAO activity was higher during the early stages of grain filling, declined thereafter and again increased towards maturity. The maximum activities of DAO coincided with the maximum content of putrescine while the levels of PAO did not seem to be directly correlated with spermidine or spermine contents. Isoelectric focusing (IEF) of DAO and PAO activities revealed the presence of bands at 30 and 45 DPA. The possible involvement of DAO and PAO in the supply of H(2)O(2) to peroxidase-catalysed reactions in the chalazal cells during grain filling is discussed.     

Diamine Oxidase Activity During the Germinative and Post-Germinative Growth of the Embryonic Axis in Chickpea Seeds

Diamine oxidase (DAO, EC 1.4.3.6.), which participates in oxidative catabolism of polyamines (PAs), was not detected in the dry viable chickpea (Cicer arietinum L.) seeds. From the time when the embryonic axis acquired an aerobic metabolism, DAO increased concomitantly with the growth of the embryonic axis and at the same time with the deterioration of the cotyledons, although in these organs the values were clearly lower than in the axis. The highest DAO activity in the embryonic axis of seedlings grown for 72 and 96 h was found in the elongation, differentiation and hypocotyl zones, while the lowest was in the apex and plumule. The absence of cotyledons promoted the early appearance of DAO in the embryonic axis. When germination occurred at supraoptimal temperatures (30 – 35 °C), DAO activity was sharply inhibited both in the cotyledons and in the embryonic axis. This inhibition was accentuated further in the presence of cyclohexylamine, an inhibitor of spermidine synthase activity, to such a degree that DAO was undetectable in the cotyledons. DAO inhibition by EGTA and the pronounced reversal induced by Ca²⁺ implies that calcium may be related to DAO activity. The presence of putrescine, spermidine and spermine in the germination medium stimulated DAO activity, although this activity was inhibited when the exogenous PA was cadaverine.     

Polyamine catabolism in dormant embryos of the spindle tree (Euonymus europaeus L.) and in dormancy break obtained after treatment with gibberellic acid

Previously we showed that dormancy break of spindle tree embryos after gibberellic acid (GA₃) treatment was followed by an increase in arginine decarboxylase (ADC) activity (Béranger-Novat N. et al., Plant Sc. 102: 139–145, 1994). These results indicated that arginine decarboxylase pathway mediate hormone-induced growth responses in spindle tree embryos. In the present investigation we show that in GA₃-treated embryos diamine oxidase (DAO) increases immediately after putrescine content and the increase in DAO activity paralleles the accumulation of putrescine at the beginning of the culture (before the visible appearance of the radicle). In this system polyamine oxidase (PAO) increases immediately after DAO activity and follows closely the increase in spermidine content. These results demonstrate a direct correlation between the biosynthesis and oxidation of putrescine and spermidine. At every stage of development DAO and putrescine levels are lower than spermidine and PAO levels. Dormant embryos can be distinguished from GA₃-treated embryos by a complete lack of putrescine accumulation. In dormant embryos compared to GA₃-treated embryos DAO changed more or less in parallel and on the whole seemed to follow the same content and distribution, but the kinetics of the activation of DAOs were different in dormant embryos with a delay of 1.5 day for the first and 1 day for the second peak. During the first days of culture at least up to 4 days the distribution of spermidine and PAO in GA₃-treated embryos followed the same pattern observed in dormant embryos, but the levels of spermidine and PAO were greatly reduced in dormant embryos. On the other hand the kinetics of the activation of PAOs were different in dormant embryos with a delay of 1 day. The results suggest that dormant embryos are deficient in their ability to synthesize polyamines efficiently and support the view that spermidine catabolism (via PAO pool) is limiting in untreated embryos during the first days of culture.