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EN
Plants are constantly exposed to a variety of stressors during their lives. One of such stressors is contamination of the environment with heavy metals. Lead is one of highly toxic metals and it significantly inhibits normal plant growth. The study aimed at assessing the degree of relieving the stress caused by 1 mM Pb(NO3)2 via different biologically active substances (AsA, GSH, PP, α-Toc, SA) on the basis of the measurement of morphological (root length, coleoptile length, fresh weight), biochemical (Pro, MDA, CAT, POX) and physiological (Chl a+b, Car) traits in 10-day leaves of spring barley of the cultivar Eunova under laboratory conditions. Pb-stress reduced the fresh weight, root length and coleoptiles of the barley tested. Lead increased lipid peroxidation and Pro content, enhanced CAT and POX activity, and significantly suppressed the photosynthetic pigments content. Among the substances used in the experiment, PP, α-Toc and GSH generally relieved the toxic effect of lead to the barley seedlings to the greatest degree.
EN
The article presents peculiarities of drought stress identification in sugar beet and the alternatives for the use of additional components of cultivation technology aimed at increasing tolerance to water deficiency at the early stages of growth and development, namely: (i) soil amendment with moisture-retaining polymers; (ii) seed treatment with a growth regulator contributing to a better formation of the root system; and (iii) foliar application of micro fertilizers. The study of the state of the plant photosynthetic apparatus was carried out with the use of the devices for measuring chlorophyll fluorescence FLORATEST, developed at the Institute of Cybernetics of the National Academy of Sciences of Ukraine. The measurements were performed according to the guidelines on Determination of the Fluorescence Induction of plant Chlorophyll: Theoretical and Practical Bases of the Method. For better representativeness of the sampling, the measurements were performed at the same time of day and with the same intensity of illumination of the plants. As a result of the studies, the ratio of variable to maximum fluorescence (Fv/Fm) of the plant photosystem obtained with the use of a portable fluorometer was found to be the most effective method of rapid diagnostics of drought stress in plants. A high level of correlation was found between the concentration of free proline and the Fv/Fm ratio, with the correlation coefficient for sugar beet r = -0.96, which corresponds to a very strong relationship.
EN
This study aimed to investigate the sugar beet genotypes for resistance to hyperthermia and obtain temperatureresistant lines. Nine hybrids and one variety of sugar beet were screened. Cotyledonary leaves and hypocotyls isolated from aseptic seedlings were used for induction of callus and subsequent subcultivation. To create hyperthermic conditions, the callus lines were maintained in thermostats at temperatures of +27 °C, +41 °C, +45 °C, and +47 °C. The effect of high temperatures on the callus tissue was assessed by the specific callus diameter index. The free proline was detected with chromatography. As result of callus tissue exposed to temperatures of +41 °C, 45 °C and 47 °C, on the 9th day of cultivation at high temperatures, significant differences were observed in the size and colouration of the callus tissues. At a moderate temperature (+41 °C), the growth of the callus mass was somewhat higher compared to the control. At a high temperature (+45 °C), the intensity of the growth processes decreased and ceased at a temperature of +47 °C. After transfer and subsequent cultivation of callus tissues in regeneration medium MSR – Murashige and Skoog medium for regeneration, all genotypes demonstrated the formation of morphological structures that initiated the formation of regenerated plants. The number of regenerated plants largely fluctuated over temperatures and almost was not related to genotypes. Consequently, the temperature-resistant lines obtained as a result of extreme heat treatment, differ in terms of the specific diameter of callus.
EN
Paronychia argentea is a wild herb plant with a high medicinal value. P. argentea plant is a neglected herb that grows without any attention in terms of research for cultivation and propagation. The conventional propagation methods of P. argentea by seeds and cutting are not preferred due to low germination percentage and cutting rooting problems. As a substitute for seed propagation, effective micropropagation protocols were developed. Using 0.6 mg/L 6-Benzylaminopurine (BAP), maximum of 3.90 (shoot/explant) was produced on Murashige and Skoog (MS) medium. In vitro growth was significantly decreased with increasing NaCl concentration. Potassium (K) and nitrogen (N) concentrations in P. argentea plantlets decreased significantly under NaCl treatment. Level of protein in leaf tissue generally decreased with increased NaCl concentrations in the medium. Proline content in P. argentea plantlets increased significantly with increasing NaCl concentrations. An increased in NaCl concentration in the medium resulted in an increase in total soluble solids (TSS) in plant tissue. Moreover, as salinity level concentration increased relative water content was decreased. High NaCl was significantly affected in vitro plants growth. P. argentea showed that in vitro P. argentea plantlets could be tolerated to in vitro salinity.
EN
In recent years there has been a dynamic development of asymmetric synthesis. Groups of researchers, particularly the one led by Benjamin List and Carlos Barbas, carried out a number of reactions and showed the effectiveness of the use of small organic molecules such as proline as catalysts. Michael addition catalyzed with proline is a particularly interesting reaction because it can be carried out in two aminocatalytic pathways. The analysis of Michael reaction reveals potential for both forms of aminocatalysis: enamine and iminium catalysis (Scheme 1) [1–14]. Presumably Michael reaction proceeds mainly according to enamine mechanism. The use of proline in Michael reaction with imine activated acceptor is slightly effective. So far the researches have shown that the modification of proline molecule or addition of other catalyst is necessary for condensation to appear. Enamine catalysis concerns the activation of carbonyl compound in situ being a donor. There is no need for enolase anion to be created earlier [2, 15–17]. When, as a result of the reaction of a,b-unsaturated carbonyl compound with proline, Michael acceptor activation appears it means that it is enamine mechanism reaction (Scheme 1) [2, 24]. One of the first examples of direct Michael reaction proceeding through enamine transition state is the reaction of cyclopentanone with nitrostyrene (Scheme 6) [20–23]. Other examples of Michael addition of ketone with nitro olefin catalysed by proline are shown in table 2 and 3 [10, 23, 30]. Nitroketones obtained in that way are useful as precursors for different organic compounds [33], also pyrrolidines [34]. Pyrrolidines are pharmacologically active and they selectively block presynaptic dopamine receptors [34] (Scheme 7). Except for Michael intermolecular reaction, intramolecular condensation adducts were also obtained. Michael intramolecular proline-catalyzed condensation in which inactive ketones transform into α,β-unsaturated carbonyl compounds was described (Scheme 9) [35, 36]. These reactions require a stoichiometric amount of a catalyst and a long time of reaction and they give as a result a little enantiomeric excess [11, 24, 35]. In 1991, Yamaguchi and co-workers carried out malonates Michael addition to α, β-unsaturated aldehydes catalyzed by L-proline [24, 39]. The reaction proceeded according to enamine mechanism, for example dimethyl malonate was reacted with hex- 2-enal in the presence of proline to give Michael adduct in 44% yield. To improve the yield an attempt of a slight modification of a proline molecule was made transforming it into proper salt. Proline lithium salt enabled to obtain the condensation product in 93% yield (Tab. 4). Regardless of a used catalyst the products in the form of racemates were obtained. In order to improve enantioselective properties of a catalyst, Michael addition of diisopropyl malonate to cycloheptenone was carried out in chloroform in the presence of different proline salts. Optimal enantioselectivity and yield was obtained by using rubidium salt (Tab. 5–7) [40, 41]. Rubidium prolinate-catalyzed Michael additions are used in industry e.g. for enantioselective synthesis of the selective serotonine reuptake inhibitior (SSRI) (–)-paroxetine (antidepressant) (Scheme 12) [24].
EN
Mannich reaction occuring among ketone, aldehyde, and amine is one of the ways of a synthesis of biologically active compounds. Reactions of this type were carried out in the presence of different catalysts [3–10], however in recent years a lot of attention has been paid to enantioselective Mannich reaction catalyzed with proline. Such reactions were carried out with the use of different compounds containing carbonyl group and the most frequently used amine was p-anisidine. The advantage of the use of p-anisidine is a possibility of conducting the direct Mannich reaction (Scheme 3). In this way β-amino ketones (Tab. 1, 2, 4) [15, 18–20, 23, 24], α-hydroxy-β-amino ketones (Tab. 3) [15, 22], and β-amino alcohols (Tab. 5, 6) [25, 26] were obtained. A possibility of syntheses of β-amino sugars and α-amino acids with their derivatives (Tab. 7) [28, 29] is worth noticing. In a great number of described reactions, the products were obtained with satisfactory yield and enantiomeric excess. Taking into consideration the difficulty of a removal of p-hydroxyphenyl group which protects amine group in the resulting products, the attempts of using different amine compounds in Mannich reactions catalyzed with proline were undertaken. The use of amines blocked by tert-butoxycarbonyl group (Boc) enabled to obtain the products with high yield and ee values (Tab. 12–15) [35–38]. However in the case of the use of Boc the reaction must be carried out in an indirect way (it is necessary to prepare imine blocked by Boc earlier).
EN
Proline in organic synthesis is used as a small molecular organocatalyst. In a catalytic act proline, similarly to an enzyme, activates reagents, stabilizes transition state and influences an orientation of substrates [1–12]. Proline works as aldolase I (so called microaldolase I). In comparison with other amino acids it shows exceptional nucleophilicity which makes imines and enamines formation easier. In the intermolecular aldol reaction proline was used for the first time by List and co-workers (Scheme 1) [3, 9, 20]. Since then an immense progress has been observed in this field. Several aldolization reactions were performed in the presence of proline. Reactions of this type proceed between the donor (nucleophile) and the acceptor (electrophile). In aldol reaction the donors can be both ketones and aldehydes which next are condensed with ketones and aldehydes acting as electrophiles (Scheme 2–18; Tab. 1–7) [21–72]. The presence of proline ensures not only high yield of homo- and heteroaldolization but mainly enables conducting enantio- and diastereoselective synthesis. Intermolecular proline-catalyzed aldol condensation proceeds according to enamine mechanism. Anti-aldols, which make a valuable source of intermediates in the synthesis of important biologically active compounds, are mainly obtained in this reaction [35–44, 54, 58, 62, 63, 68, 69, 71].
PL
Chromatografia cienkowarstwowa jest dogodną metodą do tzw. analiz przesiewowych (z ang. screen analysis) oraz szybkiego badania profili retencji z użyciem różnych faz ruchomych. Nasze badania dotyczą zastosowania termostatowanej mikrochromatografii cienkowarstwowej do optymalizacji procesu rozdzielania wybranych aminokwasów tryptofanu i proliny w układzie faz odwróconych (RP). Uzyskane dane retencyjne umożliwiły określenie optymalnych warunków rozdzielania przy zastosowaniu prostego parametru optymalizacji: maksymalnej wartości współczynnika rozdzielenia (αmax). Parametr ten był obliczany z dokładnością 2% stężenia binarnej fazy ruchomej w zakresie od 0 do 100%, na podstawie danych eksperymentalnych uzyskanych z rozdzielczością 20%. Wybrane kryterium optymalizacji umożliwia bezpośrednie przeniesienie uzyskanych wyników na systemy rozdzielania typu RP z użyciem kolumny zamiast płytek np. wysokosprawnej chromatografii kolumnowej (HPLC).
EN
Thin-layer chromatography (TLC) and high-performance thin-layer chromatography (HPTLC) are commonly used in separation, identification, purification and quantification of different analytes present in complex biological, environmental and pharmaceutical samples. Planar chromatography can be also applied to screen analysis and fast exploration of chromatographic retention profiles using different concentrations of mobile phases components. This method does not require expensive equipment, tedious and time-consuming sample pretreatments and, what is important, allows a parallel separation of many samples at the same time. The spots or bands sprayed onto the TLC plate can be easily detected under visible and UV light (usually 254nm, 366nm) or by post-chromatographic derivatization with visualization reagents, and then digitalized using office scanners. It should be noted that the great advantage of micro-planar chromatography compared to the regular sized TLC is low consumption of the eluents (usually less than 1 mL per analytical run) as well as short analysis time, due to mobile phase migration distance less than 50 mm. Under particular cases (high temperature and low viscosity eluents like acetone, dichloro-methane or n-hexane), non-forced eluent flow micro-chromatographic analysis can be completed within less than 5 minutes. This work is focused on optimization of separation process of two amino acids including tryptophan and proline (Fig. 1) under reversed-phase micro-chromatographic conditions. Particularly, selected amino acids were chromatographed on HPTLC RP-18W (wettable with water) 25 x 50 mm cutted plates using thermostated horizontal micro-chamber and unsaturated conditions (Fig. 2). The target components were sprayed on the start line (1Μg per lane) using a semi-automatic sampler (Linomat 5, Camag). The plates were developed with n-propanol: water binary mixtures as the mobile phases. Different concentrations of alcohol ranging from 0 to 100% (v/v) with steps of 20% were applied. The temperature of chromatographic separation was set at 40oC and chromatographic runs were completed within 10-30 minutes, depending on the water concentration in the mobile phase. The amino acids were visualized by dipping the plates in ninhydrin solution (prepared for concentration 0.3% in ethanol) followed by heating at 120°C for 10 minutes. Picture acquisition was performed using a Plustek OpticPro S12 scanner with an 8-bit per channel color deep mode, 300 DPI resolution, and saved as TIFF files. Retention data were derived from the plates via densitometric scans obtained with help of ImageJ freeware (http://rsb.info.nih.gov/ij). Using the raw retention data set (RF values, retardation factor), the chromatographic profiles of amino acids were calculated with step of 2% (Fig. 3A). Based on the curves observed and relationship RM = logk, where RM and k correspond to mobility and retention factors, respectively, a simple optimization criterion expressed as a separation factor (α) was calculated (Fig. 3B). Using this plot, the best separation conditions of target components were selected taking into account the maximum value of the separation factor (αmax). Due to the equal spot shape of amino acids investigated across the whole range of the mobile phases composition, such approach involving the α values allows fast optimization of separation in planar chromatography and direct application of selected conditions into column chromatographic systems based on e.g. high-performance liquid chromatography (HPLC). It should be noted that the advantages of retention data screening using micro-TLC are very low mobile phase consumption (at level of 300 ΜL per chromatographic run), short analysis time and effective detection of UV-Vis transparent substances using simple visualization reagent.
EN
The antioxidant system effects of Kandelia candel were investigated under four different levels of PAH stress. The activities of antioxidant enzymes including superoxide dismutase (SOD), catalase (CAT) and peroxidase (POD), the responses to the change of malondialdehyde (MDA) contents and the accumulation of proline in K. candel were determined. Our results suggested that the activities of SOD, CAT, POD increased significantly in leaves and roots of K. candel (p≤0.05) with the increase of the external PAH concentrations, while in stems, the activities of these antioxidant enzymes were all significantly inhibited (p≤0.01). We also observed an increase of MDA in leaves, stems and roots, and an obvious correlation between MDA content and PAH concentrations in three locations, which showed that the change of MDA content could be used as a biomarker of K. candel under PAH stress. The proline content was found remarkably enhanced in leaves, stems and roots. However, a significant inverse correlation was observed between the proline content and SOD (r=-0.99, p≤0.01), POD (r=-0.95, p≤0.05) activities in stems. This study suggested that the antioxidative system of K. candel has an obvious organ-dependent feature when exposed to PAH contamination as revealed by discriminant analysis (DA).
EN
The study aimed at demonstrating intracellular proline synthesis under osmotic stress conditions in selected soil fungi on the example of Trichoderma sp. and Trichotecium roseum. Effect of the increase of sodium chloride (NaCl) salinity, in concentrations of 1 to 1000 mmol dm–3 PDA medium, on proline content in the fresh matter of mycelium cultured on medium was examined. The increase of medium salinity affects production of osmoregulatory substances in the form of proline in selected soil fungi. Its content in mycelium depends on salt (NaCl) concentration in medium as well as on species. Intracellular proline synthesis increased starting with the least salinity. As osmotic stress increased, proline content in mycelium almost quadrupled in Trichoderma sp. and quintupled in Trichotecium roseum.
PL
Celem badań było wykazanie syntezy wewnątrzkomórkowej proliny w warunkach stresu osmotycznego u wybranych grzybów glebowych na przykładzie Trichoderma sp. i Trichotecium roseum. Badano wpływ stresu osmotycznego w wyniku zasolenia chlorkiem sodu (NaCl), w stężeniu od 1 do 1000 mmol dm-3 pożywki PDA na zawartosć proliny w świeżej masie grzybni wyrosłej na podłożu. Wzrost zasolenia podłoża wpływa na wytwarzanie substancji osmoregulacyjnych w postaci praliny u wybranych grzybów glebowych. Jej zawartość w grzybni jest zależna od stężenia soli NaCl w podłożu oraz od gatunku. Synteza proliny wewnątrzkomórkowej zwiększała się począwszy od najmniejszego zasolenia. W miarę wzrostu stresu osmotycznego zawartość proliny w grzybni zwiększyła się prawie czterokrotnie u Trichoderma sp. i pięciokrotnie Trichotecium roseum.
EN
Atmospheric ozone remains depleted which in turn leads to the increase of UV-B radiation reaching the surface of the earth and in the same time more and more nitrogen will be imported into the terrestrial ecosystems through nitrogen deposition. These two factors will operate simultaneously. The photosynthetic and physiological responses of deciduous broad leaved species Swida hemsleyi occurring commonly at 1350-3700 m a.s.l. subjected to enhanced UV-B and to nitrogen supply were studied. The experimental design included two levels of UV-B treatments (ambient UV-B, 11.02 KJ m[^-2] day[^-1] and enhanced UV-B, 14.33 KJ m[^-2] day[^-1] and two nitrogen levels (without supplemental nitrogen supply and with supplemental nitrogen supply). An experiment was conducted in open semi-field condition in Maoxian Ecological Station of Chinese Academy of Sciences, Sichuan province, China at 1820 m a.s.l. Enhanced UV-B caused a marked decline in growth parameters, net photosynthetic rate, stomatal conductance to water vapour, chlorophyll pigments, whereas it induced an increase in rate of reactive oxygen species (ROS) production and ROS accumulation and malondialdehyde (MDA) content. Enhanced UV-B also induced an increase in leaf thickness and antioxidant compounds content, such as carotenoids and proline content. On the other hand, nitrogen supply caused an increase in some growth parameters, chlorophyll pigments and antioxidant compounds, and reduced ROS accumulation. However, nitrogen supply did not affect MDA content under enhanced UV-B, though it increased antioxidant compounds content and reduced the rate of ROS production and ROS accumulation. These results implied that enhanced UV-B brought harmful effects on Swida hemsleyi seedlings and supplemental nitrogen supply could alleviate the adverse effects of UV-B radiation on plants to some extent.
EN
Androgenic embryos of 3 different carrot genotypes were cultivated on the medium containing different Cu2+ concentrations: 0.1 uM (control), l uM, 10 uM and 100 uM. Carrot sensitivity to Cu2+was evaluated on the basis of growth inhibition, changes in peroxidation of membranes and proline content. The cultivation on the medium supplemented with Cu2+ resulted in dose-dependent inhibition of the growth and organogenic ability of Narbonne carrot embryos, while in the genotype 1014 treated with 10 uM Cu2+ the significant regeneration ability was obseryed. The same Cu2+ concentration greatly increased the level of free proline in the genotype 1014 which was accompanied by relatively low TBARS content. In the genotype Feria 100 uM Cu2+ concentration triggered large increase in proline content associated with high regeneration capacity of carrot embryos.
PL
Zarodki androgenne 3 różnych odmian marchwi hodowano na pożywce z dodatkiem Cu2+ w 4 stężeniach: 0,1 uM (kontr.), l uM, 10 uM, i 100 uM. Wrażliwość marchwi na Cu2+oceniano na podstawie zahamowania wzrostu, zmian w utlenianiu lipidów błonowych oraz zawartości wolnej proliny. Hodowla na pożywce wzbogaconej w Cu2+ spowodowała stopniowe zahamowanie wzrostu oraz zdolności regeneracyjnych zarodków marchwi odm. Narbonne, podczas gdy ilość prawidłowych rozet genotypu 1014 traktowanych 10 uM Cu2+ była największa. Badany metal w tym samym stężeniu znacząco zwiększył zawartość wolnej proliny w rozetach genotypu 1014, co było skorelowane ze stosunkowo niskim poziomem TBARS. Natomiast, w przypadku genotypu Feria, wzrost zawartości proliny skojarzonej z dużymi zdolnościami regeneracyjnymi zarodków zaobserwowano w rozetach traktowanych Cu2+ o stężeniu 100 uM.
EN
A two-factor hydroponic experiment was conducted on nutrient medium whose concentrations were as follows: 0.068, 0.136 and 0.170 mol/dm3. During the experiment, the proline content was determined 12, 48 and 168 hours after applying the stress agent in the leaves of three basket willow clones using the method of BATES at al. [1]. The study aimed at determining the effect of different NaCl concentrations in substrate as well as the influence of stress time on the proline content in the leaves of basket willow clones ("Bjor", "Jorr" and "Tora"). The "Bjor" clone was found to be characterised by the highest proline content, whose concentration rose together with an increase in the salinity of nutrient medium and the time of osmotic stress applied. The "Jorr" clone was characterised by the lowest proline content in leaf fresh mass, with its maximum value being found after 48 hours of stress duration.
PL
Przeprowadzono dwuczynnikowe doświadczenie hydroponiczne, w którym stężenia pożywki wynosiły kolejno 0,068; 0,136; 0,170 mol/dm3. Oznaczono zawartość proliny metodą Batesa i wsp. [1] po 12, 48, 168 godzinach od momentu zastosowania czynnika stresowego w liściach trzech klonów wierzby wiciowej. Celem pracy było określenie wpływu zróżnicowanych stężeń NaCl w podłożu oraz czasu oddziaływania stresu na zawartość proliny w liściach klonów wierzby "Bjor", "Jorr", "Tora". Stwierdzono, że klon "Bjor" charakteryzował się największą zawartością proliny, a jej stężenie rosło w miarę zwiększania zasolenia pożywki oraz wydłużania czasu oddziaływania zastosowanego stresu. Klon "Jorr" charakteryzował się najmniejszą ilością proliny w świeżej masie liści, a jej maksymalną ilość stwierdzono po 48 godzinach oddziaływania stresu.
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