Accelerated ageing and subsequent imbibition affect seed viability and the efficiency of antioxidant system in macaw palm seeds

Accelerated ageing is an accurate test indicator of seed vigor and storability that helps to understand the mechanisms of cellular and biochemical deterioration that occur during seed ageing. This study was carried out to elucidate the mechanisms of ageing in macaw palm embryos. Seeds were artificially aged during 4, 8 and 12 days at 45 °C and 100% relative humidity. After ageing, seeds were tested for viability (tetrazolium), electrical conductivity, lipid peroxidation (MDA) and hydrogen peroxide (H₂O₂) content. Part of the aged seeds was imbibed for 8 days and then determined the hydrogen peroxide content and the activity of antioxidant system enzymes (superoxide dismutase, catalase and glutathione reductase). Ageing reduced the embryo viability from 8 days of treatment and increased malondialdehyde content (MDA) and solute leakage. Hence, membrane permeability correlated with both loss of viability and lipid peroxidation. Imbibition after ageing significantly increased H₂O₂ content along with superoxide dismutase activity. Catalase activity was significantly higher than control in embryos aged from 8 days and imbibed, and glutathione reductase activity did not change. Our results suggest that macaw palm seed deterioration during accelerated ageing is closely related to lipid peroxidation, and that enzymatic antioxidant system is not completely efficient in reducing reactive oxygen species after imbibition, a critical phase to germination. Moreover, accelerated ageing test can be used as a reliable model to understand the mechanisms involved in palm seeds deterioration.     

Ageing of isolated embryos and endosperms of durum wheat: An analysis of chromosome damage

Embryos (EM) and endosperms (END) were isolated from durum wheat seeds and aged separately for 3 years. Analysis of chromosome damage was carried out on root tip meristems of transplants in which aged (a) embryos and/or endosperms were used. The aim of the experiments was to investigate the ageing process when embryos and endosperms age separately and to evaluate the possible influence of the endosperm during storage of the seed.The results obtained indicate that (i) both aged embryo and endosperm produce mutagenic substances capable of inducing nuclear damage in the radicle meristem and (ii) age-induced damage in the embryo is not the consequence of endosperm ageing. The relationships between chromosome damage produced by aged embryos and endosperms in the transplants are discussed.     

Proteolytic Activities in x Haynaldoticum sardoum Seeds of Different Ages

Carboxypeptidase, aminopeptidase and proteinase activities weremeasured in endosperms from dry and germinating x Haynaldoticumsardoum naturally aged seeds. Carboxypeptidase activity, presentin dry seeds, decreased slightly during germination and remainednearly unchanged during the storage period. Aminopeptidase activityincreased during germination in younger seeds, but decreasedin non-viable seeds. Proteinase activity was absent in dry seeds,increased during germination in younger seeds and disappearedin the older ones. Proteinase activity was not recovered in old endosperms followingtransplantation of young embryos, and was recovered only toa very small extent in young endosperms following transplantationof old embryos. Young endosperms onto which young embryos hadbeen transplanted gave maximum recovery of enzyme activity,although this was lower than in young intact seeds. These results suggest that the reduced or delayed availabilityof nutrients to the embryo axis is not the only factor causingthe failure of seeds to germinate, the ageing process beinga progressive phenomenon affecting both embryo and endosperm. x Haynaldoticum sardoum, Denti de cani, seed ageing, proteolytic activities, embryo-transplantation     

Function of the ascorbate-glutathione cycle in aged sunflower seeds

The function of the ascorbate-glutathione (AsA/GSH) cycle was analyzed in seeds of sunflower ( Helianthus annuus L. cv. Peredovik) subjected to accelerated ageing at 43°C and 75% relative humidity for 1 to 11 days. The study was performed using dry seeds and seeds hydrated by imbibition in distilled water for 4 h at 25 °C. Lipid peroxidation was also determined by measuring the malondialdehyde (MDA) level. As the ageing period increased, a progressive loss of seed viability became increasingly evident. Even though high levels of MDA were delected, the MDA level did not change during accelerated ageing, suggesting that lipid peroxidation might occur to some extent. The study of the ascorbate/glutathione (AsA/GSH) cycle revealed that the GSH system is the major detoxifying mechanism in both dry and imbibed sunflower seeds. The GSH system is mainly located in the embryo, and its protective role is mediated by reactions that consume the GSH pool and, thereby, minimize the increase of the oxidized form (GSSG). Seed imbibition activates cellular metabolism and allows some antioxidant enzymes like glutathione reductase (EC 1,6,4,2) to act upon toxic agents. These reactions provide a reducing status, so that repair of damage becomes possible. However, prolonged ageing conditions (11 days) result in an irreversible damage, as evidenced by the appearance of dead seeds when the germination period ended. Multiple regression analysis revealed the effectiveness of the GSH system in aged seeds, especially upon imbibition and until the AsA/GSH cycle became completely functional.     

Inter-nucleosomal DNA fragmentation and loss of RNA integrity during seed ageing

The germination of viable seeds is the basis for new plant growth and development. Seeds lose viability during storage, but the biochemical mechanisms of seed death are not fully understood. This study aimed to investigate degradation patterns of nucleic acids during seed ageing and subsequent water uptake. Seeds of Pisum sativum L. were artificially aged at 50°C and 12% seed water content (WC). Nucleic acids degradation was studied during ageing and during imbibition of four seed lots with differential viability from highly viable to dead. As seeds lost viability during ageing, DNA was gradually degraded into internucleosomal fragments, resulting in ‘DNA laddering’, in conjunction with disintegration of 18S and 28S rRNA bands. During imbibition, non-aged controls had high levels of DNA and RNA integrity through to radicle protrusion. In an aged seed lot with 85% total germination (TG) DNA fragmentation decreased upon imbibition probably due to nucleosome degradation, while rRNA integrity did not improve. In an aged seed lot with 44% TG, neither DNA nor rRNA integrity improved upon imbibition. Dead seeds showed DNA degradation as laddering throughout imbibition along with extensive degradation of rRNA. We present a model in which interlinked programmed and non-programmed events contribute to seed ageing, and suggest that protection of nucleic acids during ageing is key to seed longevity.     

Induction of RNase and nuclease activity in cultured maize endosperm cells following sucrose starvation

The activity of RNases and nucleases in plants often increases following exposure to many types of stress, including prolonged exposure to dark or phosphate starvation. In cereals, the activity of RNases and nucleases is also regulated developmentally during late seed development. In this study, we investigated the effect that the absence of sugar or phosphate in culture medium has on the activity of RNases and nucleases expressed in maize endosperm suspension cells. Withdrawal of sugar from the culture medium resulted in a substantial increase in RNase and nuclease activities, whereas deprivation of phosphate during the same period of growth had no detectable effect on either of these activities. The increase in RNase activity was limited to the neutral RNases, demonstrating that the effect of sugar starvation is specific to one class of RNase. Elimination of asparagine from the medium resulted in a transient reduction in nuclease but not in RNase activity. These observations suggest that sugar starvation constitutes a stress to which maize endosperm responds, in part, by increasing neutral RNase and nuclease activity.     

Biochemical changes induced by accelerated aging in sunflower seeds. I. Lipid peroxidation and membrane damage

When stored at 42°C and 100% relative humidity for 1 to 8 days, sunflower seeds (Helianthus annuus L. cv. Rodeo) aged prematurely and lost 25% of their initial viability. A ten-fold increase in conjugated dienes as well as a decrease of unsaturated fatty acids in diacylglycerol and polar lipids fractions were observed after 8 days of accelerated aging, demonstrating the occurrence of lipid peroxidation in prematurely aged sunflower seeds. However, the viability remained relatively high. The absence of membrane damage in seeds and of lipid peroxidation in isolated microsqmes suggested that lipid peroxidation concerned mainly lipid reserves. These results suggest that, at least within the first 8 days of treatment, the lipid reserve in sunflower seeds might act as a detoxifying trap, protecting membranes from excessive damage.     

Ageing-induced changes in glutathione system of sunflower seeds

The glutathione system is thought to be involved in defence mechanisms present in plant tissues. The efficacy of this system was evaluated in large seeds of sunflower ( Helianthus annuus L. cv. Peredovik) in response to accelerated ageing (43°C/75% relative humidity from 1 to 11 days). Differences between the embryo axis and cotyledons in relation to the glutathione system were also investigated. Additionally, lipid peroxidation was determined by measuring the malondialdehyde (MDA) content. All assays were performed using dry seeds and seeds subsequently hydrated by imbibition in distilled water for 12 h at 25°C. Accelerated ageing caused a marked decrease in seed viability, accompanied by an increase in mean germination time. There were no changes in total glutathione in dry seeds. However, the distribution in its reduced (GSH) and oxidized (GSSG) forms revealed that ageing produced a slow conversion from GSH to GSSG. As the ageing period increased, this effect was accompanied by a decrease in glutathione reductase (GR, EC 1.6.4.2) activity. The results also indicated that the GSH system exerts a different response in the embryo axis as compared with the cotyledon: (1) the GSH levels decreased less in the cotyledons than in axes of aged seeds, and (2) the GSSG level in cotyledons was independent of ageing, while its amount increased in aged embryo axes. These different responses, in conjunction with the lower MDA levels in large as compared with small seeds, indicate a possible protective role of the reserve lipids. The efficacy of the GSH system in aged seeds was associated with seed viability, as revealed by multiple regression analysis. Upon imbibition, aged seeds were able to restore their GSH levels, reaching values approximating those of unaged seeds.     

The nuclear protein Poly(ADP‐ribose) polymerase 3 (AtPARP3) is required for seed storability in Arabidopsis thaliana

The deterioration of seeds during prolonged storage results in a reduction of viability and germination rate. DNA damage is one of the major cellular defects associated with seed deterioration. It is provoked by the formation of reactive oxygen species (ROS) even in the quiescent state of the desiccated seed. In contrast to other stages of seed life, DNA repair during storage is hindered through the low seed water content; thereby DNA lesions can accumulate. To allow subsequent seedling development, DNA repair has thus to be initiated immediately upon imbibition. Poly(ADP‐ribose) polymerases (PARPs) are important components in the DNA damage response in humans. Arabidopsis thaliana contains three homologues to the human HsPARP1 protein. Of these three, only AtPARP3 was very highly expressed in seeds. Histochemical GUS staining of embryos and endosperm layers revealed strong promoter activity of AtPARP3 during all steps of germination. This coincided with high ROS activity and indicated a role of the nuclear‐localised AtPARP3 in DNA repair during germination. Accordingly, stored parp3‐1 mutant seeds lacking AtPARP3 expression displayed a delay in germination as compared to Col‐0 wild‐type seeds. A controlled deterioration test showed that the mutant seeds were hypersensitive to unfavourable storage conditions. The results demonstrate that AtPARP3 is an important component of seed storability and viability.     

Effects of storage temperature on viability, germination and antioxidant metabolism in Ginkgo biloba L. seeds.

The behaviour of the Ginkgo biloba L. seeds was studied during storage at 4 and 25 degrees C. When stored at 25 degrees C, all the seeds died in 6 months. Cold temperatures preserved seed tissue viability for 1 year but did not preserve their capability to germinate, since such capability decreased after 6 months. A significant increase in lipid peroxidation occurred in the seed both in the embryo and in the endosperm. During storage a progressive deterioration of the endosperm tissues was evident. The two major water soluble antioxidants, ascorbate (ASC) and glutathione (GSH), showed different behaviour in the two conditions of storage and in the two main structures of the seed, the embryo and the endosperm. The ASC content of embryos and endosperms remained quite unchanged in the first 9 months at 4 degrees C, then increased. At 25 degrees C a significant decrease in the ASC content in the embryos was evident, whereas it remained more stable in the endosperm. The GSH pool decreased at both storage temperatures in the embryos. As far as the ASC-GSH redox enzymes are concerned, their activities decreased with storage, but changes appeared to be time-dependent more than temperature-dependent, with the exception of the endosperm ascorbate free radical (AFR) reductase (EC 1.6.5.4), the activity of which rapidly decreased at 25 degrees C. Therefore overall the antioxidant enzymes were scarcely regulated and unable to counteract oxidative stress occurring during the long-term storage.     

Content and De Novo Synthesis of Cocaine in Embryos and Endosperms from Fruit of Erythroxylum coca Lam

There is controversy as to whether the mature fruit of Erythroxylumcoca var. coca Lam. contains the cocaine alkaloid (benzoylmethylecgonine).In the present study, cocaine was monitored to determine ifit was present in embryos and endosperms of mature fruit ofE. coca var. coca Lam., and if present, the time required forde novo synthesis in imbibing seed. Seeds from mature fruitof E. coca were dissected to separate the embryos from the endosperms.The separated embryos and endosperms were analysed for cocaine.Subsequently, endosperms and embryos from seed imbibed. undera light and dark treatment were separated on days 3, 6, 9, 12and 15 and analysed for cocaine. Cocaine was present in embryos(0.005% of d. wt) and endosperms (0–001% of d. wt) ofmature fruit of E. coca. De novo synthesis of cocaine occurredonly in embryos of seed imbibed under light after day 9 of imbibition. Erythroxylum coca, alkaloid, benzoylmethylecgonine, cocaine, embryo, endosperm, seed imbibition     

Mechanisms of seed ageing under different storage conditions for Vigna radiata (L.) Wilczek: lipid peroxidation,sugar hydrolysis,Maillard reactions and their relationship to glass state transition

Two primary biochemical reactions in seed ageing (lipid peroxidation and non-enzymatic protein glycosylation with reducing sugars) have been studied under different seed water contents and storage temperatures, and the role of the glassy state in retarding biochemical deterioration examined. The viability loss of Vigna radiata seeds during storage is associated with Maillard reactions; however, the contribution of primary biochemical reactions varies under different storage conditions. Biochemical deterioration and viability loss are greatly retarded in seeds stored below a high critical temperature (approximately 40 degrees C above glass transition temperature). This high critical temperature corresponds to the cross-over temperature (T(c)) of glass transition where molecular dynamics changes from a solid-like system to a normal liquid system. The data show that seed ageing slows down significantly, even before seed tissue enters into the glassy state.     

大麦种子前期处理对其外植体性状的调控和对其机制的探讨(Ⅰ)

研究了种壳对大麦种子吸胀生理影响的调节作用 ,同时对其外植体性状的调控作用和其机制也进行了探讨。结果如下 :(1 )种壳显著延缓种子的吸水进程 ;(2 )种壳对分子渗漏影响显著 ;(3 )总 N的渗漏主要来自胚乳 ,Mn+ + 的渗漏在胚中相对较多 ,种壳对 K+ 渗漏影响较大 ;(4 )去壳促进吸胀期间的可溶性蛋白量和过氧化物酶同工酶活性上升 ;(5 )吸胀使 IAA、GA3含量上升 ,使胚乳中的 ABA含量显著减少 ,去壳显著促进吸胀初期胚乳中的 ABA、IAA、GA3含量上升 ,去壳减少吸胀后期胚中的 IAA、ABA、GA3含量 ;(6)去壳减少吸胀 2 4h并且种子进行培养时的培养效果 ,它们具有明显较高的萌发率和愈伤组织诱导率。     

Seed ageing-induced inhibition of germination and post-germination root growth is related to lower activity of plasma membrane H(+)-ATPase in maize roots

Seeds of most crops can be severely damaged and lose vigor when stored under conditions of high humidity and temperature. The aged seeds are characterized by delayed germination and slow post-germination growth. To date, little is known about the physiological mechanisms responsible for slow root growth of seedlings derived from aged seeds. Plasma membrane H(+)-ATPase is a universal H(+) pump in plant cells and is involved in various physiological processes including the elongation growth of plant cells. In the present study, we investigated the effect of a mild seed ageing treatment on plasma membrane H(+)-ATPase activity of seedling roots. Maize (Zea mays L.) seeds with 17% water content were aged at 45 degrees C for 30h. The aged seeds showed a 20% reduction in germination. Seedlings from aged seeds grew slowly during an experimental period of 120h after imbibition. Plasma membranes of maize seedling roots were isolated for investigation in vitro. Plasma membrane H(+)-ATPase (EC 3.6.3.6) activity was 14% lower for seedling roots developed from aged seeds as compared to control seeds. Protein gel immunoblotting analysis demonstrated that the reduced activity of plasma membrane H(+)-ATPase was attributed to a decrease in steady-state protein concentration of this enzyme. In conclusion, seed ageing causes a lower steady-state enzyme concentration of the H(+)-ATPase in the plasma membrane, which is related to slow germination and post-germination growth of seedling roots.     

An interpretation of genomic balance in seed formation in interspecific hybrids of Solanum

Summary To investigate the mechanisms of seed failure in intraspecific and interspecific crosses of Solanum two diploid, S. commersonii and Group Phureja, and one tetraploid species, S. acaule, species were crossed and the seeds were analyzed for embryo and endosperm development. Many seeds of certain crosses observed seven days after pollinations were found to contain abnormal embryos and degenerating endosperms. In some cases seeds contained an embryo with no endosperm, or an endosperm with no embryo. Other interspecific crosses which were predicted to fail actually produced seeds with normally developed embryos and endosperms. To further characterize the intra- and interspecific embryos and endosperms the nuclear DNA was measured. There are several ways to explain the ploidy levels of embryos and endosperms among the crosses, the occurrence of unreduced gametes in some cases and genomic instability in other cases. The latter resulted in chromosome loss at meiotic and mitotic divisions. Genomic balance in interspecific seeds is critical to both embryo and endosperm development.Scientific Journal Series Article No. 14636 of the Minnesota Experiment Station     

Lipid peroxidation associated with accelerated aging of soybean axes

Soybean seeds age rapidly during storage at high temperature and high relative humidity. The axes of such aged seeds contain high levels of malondialdehyde, a product of the peroxidation of unsaturated fatty acids. The levels of linoleic (18:2) and linolenic (18:3) acids in a polar lipid (phospholipid) fraction decrease during aging and more dramatically during postaging deterioration. None of these changes occurred in seeds that have been stored at high temperature but low relative humidity. No superoxide dismutase activity was detected in any nonimbibed seed. In viable seeds, activity was detectable 1.5 hours after the onset of imbibition, but none was found in aged seeds up to 5 hours. It is suggested that aging leads to peroxidative changes to lipids and that these could contribute to loss of viability.     

Biochemical changes induced by accelerated ageing in Bambusa bambos seeds

Decrease in seed viability and germination rate may be caused by biochemical changes associated with seed ageing. Different biochemical assays were conducted to investigate the changes occurring at the ageing of Bambusa bambos seeds. A reduction in the total content of food reserves such as sugars, proteins and lipids were recorded. Decreased activity of peroxidase, acid phosphatase, alkaline phosphatase were also noticed during accelerated ageing. A substantial increase in total free amino acids and the activity of amylases confirms the degradation of stored biomolecules in seeds during ageing. This revised version was published online in August 2006 with corrections to the Cover Date.     

Possible involvement of aquaporins in water uptake by pea seeds differing in quality

Using the method of room temperature phosphorescence (RTP), we divided air-dry pea (Pisum sativum L.) seeds subjected to accelerated ageing (40°C, 85% relative humidity) into three fractions: (I) high-quality seeds, (II) weakened seeds, and (III) dead seeds. In the process of ageing, seed germinability firstly decreased and then increased due to so-called “improved” seeds of fraction II, which returned to fraction I as judged from the RTP level; the germinability of these seeds became equal to that of fraction I seeds. Seeds capable of germination (fractions I and II) differed in the rates of imbibition, which depended on plasma membrane permeability (opened or closed water channels) but not on the presence of the seed coat. A low activation energy of seed imbibition in fraction II (less than 5 kcal/mol) indicates that water channels are open. A mercury-containing compound (5 μM p-chloromercuribenzoate (PCMB) reduced the rate of water uptake by these seeds, and dithiothreitol restored it. A high activation energy of fraction I seed imbibition (more than 12 kcal/mol) corresponded to the water uptake mainly across the lipid bilayer when water channels are closed. PCMB did not affect the rate of fraction I seed imbibition. We supposed that mature air-dry pea seeds had open water channels. During the first stages of fraction I seed imbibition, these channels were closed, limiting water uptake. NaF (100 μM), an inhibitor of phosphatase, prevented channel closing and accelerated the imbibition of fraction I seeds. It did not affect the imbibition rate of fraction II seeds, indicating their water channels to be opened. However, NaF did not affect the water uptake of “improved” fraction II seeds as well. It seems likely that their channels were closed during accelerated ageing but otherwise than via dephosphorylation. The results obtained indicate the possibility of water inflow regulation in the weakened seeds via the state of aquaporins, which form water channels in the membranes.     

Improving germination and vigour of aged and stored onion seeds by matriconditioning

Germination and vigour of accelerated aged (AA) and naturally stored onion seeds were examined. Accelerated ageing was conducted at 40 °C and 100 % relative humidity (RH). Non aged seeds were stored for 34 months at 3 or 15 °C and 40, 60 or 90 % RH. To restore seed viability, stored and aged seeds were matriconditioned with Micro-Cel E. A distinct loss of germination was observed after 5 days of accelerated ageing. Naturally stored seeds maintained high viability for 34 months, when stored at 3 °C and 40, 60 and 90 % RH or at 15 °C and 40 %. An increase of RH to 60 and 90 % at 15 °C caused loss of germination and vigour. Matriconditioning improved germination and increased endogenic ethylene release and in vivo ACC oxidase activity of both aged and stored seeds.