Browsing by Subject "anthocyanin"

Climate warming is expected to cause changes in winter conditions in northern regions. These changes include reduced depth and duration of the snow cover, and strong fluctuations in winter temperatures. A mesocosm experiment was planned to study the short term effects of contrasting winter conditions, and an introduced species (garden lupin; Lupinus polyphyllus), on chlorophyll fluorescence and pigment concentrations of native meadow species in southern Finland. Twelve different meadow species, representing different overwintering strategies were planted in each mesocosm at the beginning of summer in 2016 in Viikki, Helsinki. One year later, a lupin was planted in half of the mesocosms. Over the winter 2017-18, one half of the mesocosms was moved to Nåtö on the Åland islands, and the other half was moved to Lammi, Hämeenlinna. To each site, both lupin-containing mesocosms and lupin free controls were moved. In the inland site in Lammi, the mesocosms spent the winter covered by a thick snow cover that isolated them from harsh air temperatures from beginning of December to end of March. In coastal Nåtö, a thin snow cover formed in January and melted by mid-March. In the experiment, the maritime winter climate on Nåtö represented such winter conditions that are expected to be common on the mainland in the future, when climate warming progresses. Leaf chlorophyll fluorescence as well as concentrations of leaf chlorophyll and flavonoids were repeatedly measured nondestructively for all species using optical apparatus. Growth and flowering of the lupin was monitored during spring and summer 2018. No marked differences were observed in the meadow species chlorophyll fluorescence and content between sites, indicating that these are well adapted to variable winter conditions. The flavonoid composition of the meadow species seemed to be regulated by seasonal changes in light intensity and temperature. Small reductions in chlorophyll content for some species indicated that these were disadvantaged by the lupins presence. This was attributed to the lupins shadowing effect. In contrast, two evergreen species seemed to take advantage of the nitrogen input from the lupin in terms of higher chlorophyll content in summer 2018. The lupin overwintered successfully in mainland Lammi, but seemed to suffer from the maritime and snow poor winter conditions in Nåtö, which led to reduced production of leaves and inflorescences during the growing season 2018. The results indicate that native meadow species in Finland are relatively tolerant of the expected changes in mainland winter conditions, whereas these changes will be disadvantageous for the lupin.

Punaisen värin ruskistuminen on laatuongelma mansikkasoseessa. Mansikan punainen väri on peräisin antosyaaneista, jotka ovat reaktiivisia ja hajoamisherkkiä yhdisteitä. Ruskistumisreaktiot jaotellaan entsymaattisiin ja ei-entsymaattisiin reaktioihin. Kirjallisuuskatsauksessa perehdyttiin mansikan antosyaanien kemiaan ja kartoitettiin mansikkasoseen ruskistumisen eri mekanismit. Kokeellisen tutkimuksen tavoitteena oli selvittää hallintakeinoja mansikkasoseen ruskistumisen estämiseksi. Kokeellinen tutkimus koostui kolmesta osasta: kuumennus-, säilyvyys- ja prosessikokeesta. Kuumennuskokeessa selvitettiin mansikkasoseen kuumennusajan ja värimuutosten välistä yhteyttä sekä tehtiin mansikkaraaka-aineiden vertailu. Säilyvyyskokeen mansikkasosenäytteitä seurattiin kuuden viikon ajan 7 tai 23 C:n lämpötilassa. Prosessikokeessa vertailtiin teollisessa mittakaavassa valmistettujen vadelma- ja mansikkasoseiden värimuutoksia kuukauden ajan. Lisäksi verrattiin kuumennus- ja korkeapainepastörointikäsittelyn vaikutusta mansikan värin pysyvyyteen. Kolorimetrillä tehdyistä värimittauksista määritettiin näytteiden väliset värimuutokset (E*). Värimittauksen lisäksi kaikista mansikkasoseista määritettiin suuntaa antava antosyaanipitoisuus spektrofotometrisellä menetelmällä. Säilyvyyskokeen mansikkasosenäytteiden tarkat antosyaanipitoisuudet ja fenolisten yhdisteiden pitoisuudet määritettiin erittäin suuren erotuskyvyn nestekromatografisella menetelmällä. Lyhyt kuumennusaika (30 s) aiheutti mansikkasoseessa suurimmat säilytyksen aikaiset värimuutokset. Pidemmälla kuumennusajalla (180 s) värimuutokset olivat suurempia kuumennuksen aikana, mutta säilytyksen aikaiset muutokset jäivät pienemmiksi. Kylmäsäilytetty mansikkasose oli näytteistä ainoa, joka ei menettänyt hyväksyttävyyttään säilyvyyskokeen aikana. Prosessikokeessa vadelmasoseen väri osoittautui merkittävästi mansikkasoseen väriä pysyvämmäksi. Korkeapainepastöroidun mansikkasoseen väri oli yhtä hyväksyttävä kuin kuumennetun, mutta rakenteen hyväksyttävyys heikkeni kuukauden säilytyksen aikana. Tulosten perusteella voidaan päätellä, että lyhyt kuumennusaika ei ollut riittävä mansikan endogeenisen entsyymiaktiivisuuden inaktivoimiseksi. Säilyvyyskokeessa ilmeni mansikan antosyaanien suuri hajoamisherkkyys, jota voidaan hillitä vain kylmäsäilytyksellä. Mansikkasoseen kylmäsäilytyksen tarpeellisuus varmistui prosessikokeessa.

Anthocyanins are an important class of flavonoids under the class of phenolic compounds and contribute to flower color variation. Gerbera hybrida is a flowering plant of Asteraceae family having mainly two colors of flowers – orange and red. Dihydroflavonol 4-reductase (DFR) is a key enzyme catalyzing a reaction in anthocyanin biosynthesis, the reduction of dihydroflavonols to leucoanthocyanidins. GDFR1-2 and GDFR1-3 are two allelic forms of gerbera DFR differing in substrate specificity for the dihydroflavonols - dihydrokaempferol, dihydroquercetin and dihydromyricetin and also differ in 13 amino acids where eight are considered to be important for substrate specificity. GDFR1-2 has strong preference for dihydrokaempferol and GDFR1-3 doesn’t have any preference for the three substrates. In order to find out the amino acids responsible for substrate specificity, swap mutations were generated between GDFR1-2 and GDFR1-3 by two PCR methods– first, running separate PCR from the templates of GDFR1-2 and GDFR1-3, making a heteroduplex by mixing separate PCR where non-matching nucleotides are expected to be corrected by E. coli and, second, by running PCR from mixed templates with short extension time of PCR to make swaps by template switching. The second method was found more effective than the first method. 81 lines (named GDAT1-81) were sequenced and 35 unique swap mutants were found. In this work the DFR assay was done from six randomly picked GDAT lines where GDAT5 had a swap in one amino acid showing still a similar pattern of substrate specificity as the reference (GDFR1-3) indicating that the mutated amino acid doesn’t have any role in substrate specificity. GDAT14 had an extra mutation (S167P) along with 2 swaps showing incapability of reducing dihydrokaempferol, demonstrating that the mutated amino acids are important and other 4 lines were identical to either GDFR1-2 or to GDFR1-3. This was a preliminary test with 6 lines. In order to get more explanations about the roles of amino acids in substrate specificity, DFR assay was done for all the 81 lines in experiments outside of this thesis and five patterns of substrate specificity were identified indicating that substrate specificity of DFR can be altered by changing only three important amino acids. The amino acids at the position 85,135 and 181 in DFR coding sequence have been identified having important roles in substrate specificity. In addition, the amino acid at position 167 may have a function in making the gerbera DFR able to reduce dihydrokaempferol.

Ecophysiology and ecology in plants are strongly affected by the conditions surrounding them. Adaptation aids plants to survive and to succeed in the prevailing conditions. Winter is a challenge to plants, particularly in northern latitudes and higher altitudes, because it exposes plants to cold and drought, for example. Plants survive from winter on species level with the help of genetic adaptations and as individuals also with the help of acclimation. Woodland strawberry (Fragaria vesca) has been observed to grow separate winter leaves. This allows it to continue photosynthesis in mild conditions during winter, thus improving its energy balance, and to start growing earlier than other species in the spring, which is beneficial in interspecific competition. Fragaria vesca is a species that has wide distribution in the northern hemisphere, and its genotypes are found from very different locations and conditions. However, adaptive traits such as producing a new set of leaves for winter can turn out to be a disadvantage if environmental conditions change rapidly. Climate change brings about changes that are difficult to predict, and these changes are advancing at a fast pace when compared to the developmental history of plants. The aim of this thesis was to study the effect of temperature on summer and winter leaf development, stolon formation and summer and winter leaf chlorophyll, flavonol and anthocyanin content in different Fragaria vesca genotypes. Leaf chlorophyll and secondary compound content give information about leaf development and stress reactions in plants. Plants are known to produce anthocyanins in order to protect the photosynthetic apparatus during chlorophyll recovery in leaf senescence. Anthocyanins are also produced as a response to low temperatures. Research increases knowledge of the ecophysiological and winter ecology-related processes in Fragaria vesca and in the commercially valuable Rosacea-family as well as provides information about the possible responses of these organisms to climate change. Material for the study consisted of twelve European Fragaria vesca genotypes, which had originally been collected from five countries: Norway, Finland, Germany, Italy and Spain. The genotypes had been collected from different latitudes, and they also expressed altitudinal differences. In this study, these genotypes were kept in two temperature treatments, warm (+16°C) and cold (+11°C/six weeks, after which +6°C/four weeks) at a greenhouse. Leaf development was studied by measuring summer and winter leaf middle leaflet width and length, and petiole length. Stolons from each plant individual were counted on a weekly basis and observations about stolon production in relation to the timing of summer leaf senescence and winter leaf development were made at the same time. Leaf chlorophyll and secondary compound content was measured with a Dualex-meter, which provided values for chlorophyll, flavonol and anthocyanin content. The underlying assumption was that cold temperature would induce winter leaf development and summer leaf senescence. The results show that there were differences in summer leaf size between genotypes. Winter leaves had differences between genotypes, but also within genotypes at different temperature treatments. Stolon count was lower and stolon production ceased slightly earlier in the cold treatment. Moreover, summer leaf chlorophyll content decreased in both treatments, but the summer leaves senesced earlier in the warm room. Summer leaf flavonol and anthocyanin values were generally higher in the cooler temperature treatment. Anthocyanins were also produced by winter leaves in the cooler temperature treatment. Based on the results, Fragaria vesca genotypes had differences related to their origin, but temperature also had an effect on winter leaf development, stolon production and the production of secondary compounds. The effect of cold temperature on the size of developing winter leaves was clear. In the cooler temperature treatment, the winter leaves were smaller than in the warmer treatment. The anthocyanin content of summer leaves was higher than in the winter leaves, and the summer leaf anthocyanin content was higher in the colder temperature treatment, where the stress related to the photosynthetic apparatus and low temperatures was combined. Nevertheless, lower temperature did not explain all the responses observed in the genotypes of the study, and thus it is likely that acclimation and winter leaf development in Fragaria vesca are affected by some other factor in addition to temperature, e.g. light regime. A possible continuation for this work would be to study the effect of light conditions or milder winters on winter leaf development in Fragaria vesca genotypes and on the physiology of the species.