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HPLC coupled to a mass spectrometer (MS) was used for the analysis of galanthamine and lycorine in natural extracts of Leucojum aestivum and in their in vitro cultures grown with a precursor (ACC), inhibitors (AgNO(3), STS), or an absorber (KMnO(4)) of ethylene. The maximum galanthamine (0.002%) and lycorine (0.02%) concentrations in tissue cultures were obtained in the presence of KMnO(4). GCMS was used to investigate underivatized alkaloid mixtures from L. aestivum. Seven alkaloids were identified in in vivo bulbs. KMnO(4) led to the highest diversity of alkaloids in tissue culture extracts.

Vigor and selected physiological parameters (content of phenolic compounds, soluble sugars, chlorophyll a and b, and carotenoids) of eight naked and two husked oat cultivars harvested at 15% moisture content were determined. Oat seeds were threshed using two rotational speeds of the threshing drum: 1.6 ms −1 (LS) and 2.4 ms −1 (HS). They were then inoculated with a medium pathogenicity strain of Fusarium culmorum, strain IPO 348-01. In naked cultivars, the use of HS resulted in more severe mechanical damage; in consequence, seedling vigor decreased by 16%. In naked cultivars chlorophyll a and b and carotenoids content were significantly reduced-by more than 64%-when the HS was used. The inoculation caused over a 100% increase of carbohydrates in roots at LS but only a slight increase at HS. Phenolic compound content was twice as high in roots than in leaves after inoculation for both LS and HS. Area of microdamage and reduction of root fresh weight (f.wt.) are significantly correlated with biochemical parameters. Smaller microdamage area and root f.wt. reduction are connected with higher physiological parameters, which confirms lower seedling susceptibility to pathogen infection.

Previously, our in silico analyses identified four candidate genes that might be involved in uptake and/or accumulation of arsenics in plants: arsenate reductase 2 (ACR2), phytochelatin synthase 1 (PCS1) and two multi-drug resistant proteins (MRP1 and MRP2) [Lund et al. (2010) J Biol Syst 18:223-224]. We also postulated that one of these four genes, ACR2, seems to play a central role in this process. To investigate further, we have constructed a 3D structure of the Arabidopsis thaliana ACR2 protein using the iterative implementation of the threading assembly refinement (I-TASSER) server. These analyses revealed that, for catalytic metabolism of arsenate, the arsenate binding-loop (AB-loop) and residues Phe-53, Phe-54, Cys-134, Cys-136, Cys-141, Cys-145, and Lys-135 are essential for reducing arsenate to arsenic intermediates (arsenylated enzyme-substrate intermediates) and arsenite in plants. Thus, functional predictions suggest that the ACR2 protein is involved in the conversion of arsenate to arsenite in plant cells. To validate the in silico results, we exposed a transfer-DNA (T-DNA)-tagged mutant of A. thaliana (mutation in the ACR2 gene) to various amounts of arsenic. Reverse transcriptase PCR revealed that the mutant exhibits significantly reduced expression of the ACR2 gene. Spectrophotometric analyses revealed that the amount of accumulated arsenic compounds in this mutant was approximately six times higher than that observed in control plants. The results obtained from in silico analyses are in complete agreement with those obtained in laboratory experiments.

This paper reports a continuation of our previous research on the phytochelatin synthase1 (PCS1) gene involved in binding and sequestration of heavy metals or metalloids in plant cells. Construction of a 3D structure of the Arabidopsis thaliana PCS1 protein and prediction of gene function by employing iterative implementation of the threading assembly refinement (I-TASSER) revealed that PC ligands (3GC-gamma-glutamylcysteine) and Gln50, Pro53, Ala54, Tyr55, Cys56, Ile102, Gly161, His162, Phe163, Asp204 and Arg211 residues are essential for formation of chelating complex with cadmium (Cd²⁺) or arsenite (AsIII). This finding suggests that the PCS1 protein might be involved in the production of the enzyme phytochelatin synthase, which might in turn bind, localize, store or sequester heavy metals in plant cells. For validation of the in silico results, we included a T-DNA tagged mutant of Arabidopsis thaliana, SAIL_650_C12, (mutation in AtPCS1 gene) in our investigation. Furthermore, using reverse transcriptase PCR we confirmed that the mutant does not express the AtPCS1 gene. Mutant plants of SAIL_650_C12 were exposed to various amounts of cadmium (Cd²⁺) and arsenite (AsIII) and the accumulation of these toxic metals in the plant cells was quantified spectrophotometrically. The levels of Cd²⁺ and AsIII accumulation in the mutant were approximately 2.8 and 1.6 times higher, respectively, than that observed in the wild-type controlled plants. We confirmed that the results obtained in in silico analyses complement those obtained in in vivo experiments.

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Changes in the germinability and vigour of seeds with sprouting damage obtained after induced sprouting under laboratory conditions were investigated in winter triticale cultivars: Bogo, Moreno, Vero, in the years 1998-2000. The seeds were tested directly after harvest, at the optimal time for sowing winter triticale and after accelerated ageing test. Average germinability evaluated directly after induced sprouting and at the optimal time for sowing winter triticale ranged from 78.0% to 92.7% and from 70.3% to 81.0%, respectively. Similar results were obtained for the length of the first leaf and the length of the longest root. The estimated components of variance showed that the year variation and interaction of years with seed sprouting damage made the greatest percentage (to 49.2% and to 31.8%, respectively) of the total variation. After accelerated ageing a significant decrease in germinability and vigour of seeds was observed. That decrease was in proportion to sprouting damage which accounted for 57.6% of the total variation. These results point to poor storability of seeds with sprouting damage.Keywords: winter triticale; sprouting; seed germinability and vigourThe germinability and vigour of seeds are conditioned by genetic traits, which are to a considerable degree modified by agroecological factors affecting the parent plant. After harvest, changes in germinability and vigour can be influenced by the method of storing and ennobling the seeds, however they will depend largely on the initial vigour. In the case of triticale, which displays a considerably strong tendency to sprout, conditioned by high alpha-amylase activity (Masojć and LarsonRaźnikiewicz 1991) and light dormancy (Moś 1994), the initial vigour of seeds can vary, depending on weather conditions during maturation and harvest. According to Belderok (1968), sprouting reduces the quality of seeds and their vigour. The aim of the research was to determine the effect of various sprouting damages of winter triticale seeds on changes in germinability and vigour estimated at various dates. The accelerated ageing test was used to indicate the effect of sprouting on vigour and to determine the storability of seeds with sprouting damage. MATERIAL AND METHODSThe material for the studies was obtained from observational field experiments carried out in the years 1997-1999 at the Experimental Station in Prusy near Cracow. The testing included three winter triticale cultivars: Bogo, developed at ZDHAR Małyszyn, Moreno and Vero, developed at ZHR Choryń. Spikes were harvested after they had attained full maturity. The spikes were threshed on the day of harvest.Sprouting of seeds was induced directly after harvest and threshing, under laboratory conditions, on Petri dishes, at the temperature of 20°C. Four degrees of seed sprouting damage were distinguished: seeds with the testa broken near the embryo (1 st ), seeds with visible primordia of a shoot and an embryonal root (2 nd ), seeds which developed a shoot and one root (3 rd ), and seeds which devel...

The imbibition of triticale kernels with a different degree of sprouting and with no visible sprouting symptoms was studied using digital image analysis and the classical weight method. Measurements were performed at two-hour intervals for the first 14 hours of imbibition, and at 24 and 26 hours. The area, perimeter, length and width of kernel images, and ten different shape factors were determined along with the weight of imbibing kernels. The germination capacity was determined on the seeds used for measurements. Kernels showing the lowest degree of sprouting were characterized by the highest rate of water uptake in comparison with the control. Most of shape descriptors were significantly correlated with kernel weight, especially image area (r = 0.688) and two shape factors (calculated on the basis of image perimeter and area) of S₉ and S₁₀ (r = 0.742 and 0.958, respectively), which makes them the best descriptors of image of imbibing seeds.