The foci of this dissertation are on advanced liquid chromatography (LC) separation and mass spectrometry (MS) techniques for the analysis of small bioactive molecules. In addition to discussing general aspects of such techniques the results from analyses of polyphenols (PPs), alkaloids and amino acids published in five appended studies are presented and discussed. High efficiency and well understood principles make LC the method of choice for separating analytes in many kinds of scientific investigations. Moreover, when LC is coupled to an MS instrument, analytes are separated in two stages: firstly they are separated and pre-concentrated in narrow bands using LC and then separated according to their mass-to-charge (m/z) ratios in the MS instrument. Some MS instruments can provide highly accurate molecular weight measurements and mass resolution allowing identification of unknown compounds based purely on MS data, thus making prior separation unnecessary. However, prior separation is essential for analyzing substances in most complex matrices – especially useful is the ultra-high performance LC (UHPLC). The advantages of using UHPLC rather than HPLC for the analysis of PPs in tea and wine were evaluated in one of the studies this thesis is based upon. The phenolic composition of red wine was also examined, using a novel LDI technique, following solid phase extraction (SPE). A class of small aromatic molecules (medicinally important alkaloids) also proved to be amenable to straightforward analysis, by thin layer chromatography (TLC) work-up followed by LDI-MS. Finally, a LC-MS method for monitoring neurotoxins (β-N-methyl-amino-L-alanine and 2,3-diaminobutyric acid) in complex biological matrices was developed and applied. Overall, the studies show that careful attention to the physicochemical properties of analytes can provide insights that can greatly facilitate the development of alternative methods to analyze them, e.g. by LDI. / <p>At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 4: In press. Paper 5: Manuscript.</p>

Tyrimo tikslas - pagaminti vandenines ir etanolines ištraukas iš Junglans nigra L. lapų ir įvertinti bendrą fenolinių junginių, bendrą flavonoidų, juglono kiekį bei nustatyti antioksidacinį aktyvumą. Metodika: 1. Tyrimo objektai: 70% etanolinė juodojo riešutmedžio tinktūra, vandeninė juodojo riešutmedžio lapų ištrauka bei gamyklinis “Black Walnut” 1:1 ekstraktas (gamintojas „Nature's Answer”, JAV). 2. Ištraukų iš J. nigra lapų gamyba. 1) Etanolinės tinktūros gamyba. Pritaikius perkoliacijos metodą iš juodojo riešutmedžio lapų žaliavos, naudojant 70% etanolį, buvo pagaminta tinktūra (santykis 1:10). 2) Vandeninė ištrauka. 10 g smulkintos lapų žaliavos buvo užpilta 120 ml verdančio vandens ir kaitinta 1 val. Ištrauka nufiltruota pro 4 nr. Vatmano filtrą. 3. Kokybinės flavonoidų įvertinimo reakcijos. 4. Bendras fenolinių junginių nustatymas naudojant Folin – Ciocalteu metodą (standartas – galo r.) 5. Bendras flavonoidų kiekio nustatymas spektrofotometriniu būdu, naudojant AlCl3 tirpalą (standartas - rutinas). 6. Juglono kiekio nustatymas spektrofotometriniu būdu. 7. Antioksidacinio aktyvumo įvertinimas fluorimetriniu būdu, naudojant 10-acetil-3,7-dihidroksifenoksazinas (AMR) ir krienų peroksidazę (HPR). Rezultatai. Bendram fenolinių junginių, bendram flavonoidų bei juglono išekstrahuoto kiekio palyginimui buvo pagamintos J. nigra lapų etanolinė ir vandeninė ištraukos. Tiriamų objektų biologiškai aktyvių komponentų kiekiai buvo palyginti su gamykliniu „Black Walnut“ ekstraktu... [toliau žr. visą tekstą] / Aim of the work - produce aqueous and ethanolic extracts of Junglans nigra leaves, evaluate the total phenolic and total flavonoids content, juglone concentration and identify antioxidant activity. Methods: 1. Objects of study: ethanolic tincture and aqueous extract of black walnut leaves, “Black Walnut” extract, 1:1 („Nature's Answer”, USA). 2. Liquid preparations of J. nigra leaves production: 1) Ethanolic tincture production. Tincture was produced of black walnut leaves applying the method of percolation. The 70% ethanol was used. Tincture ratio is 1:10. 2) Aqueous exract production. 10 g shredded leaves of black walnut were extracted with 120 ml of boiling wather for 1 hours. After boiling extract was filtered throught the Whatman paper no. 4. 3. Qualitative evaluation of flavonoids. 4. Total phenolic content was determined by spectrophotometric (Folin - Ciocalteau) method. 5. Total flavonoids content was measured with an aluminum chloride colorimetric assay using spectrophotometer. 6. Juglone determination by spectrophotometric method. 7. Fluorimetric antioxidant activity assay by using N-Acetyl-3,7-dihydroxyphenoxazine (Amplex Red) and horseradish peroxidase (HPR). Results. Ethanolic and aqueous extracts of J. nigra leaves were made to compare total phenolic, total flavonoids content, juglone concentration. Biologically active components study results of ethanolic and aqueous extracts were compared with “Black Walnut” extract. The highest total phenolic and total... [to full text]

Darbo tikslas: atlikti Lietuvoje registruotų mėlynių uogų pasirinktų preparatų sudėčių ir technologijos palyginamąją analizę, įvertinti kiekybiškai ir nustatyti antioksidacinį aktyvumą. Darbo uždaviniai: Ištirti Lietuvoje registruotų mėlynių uogų preparatų pasiūlą. Atlikti šių preparatų palyginamąją analizę vertinant sudėtis. Pasirinktuose preparatuose nustatyti fenolinių junginių ir antocianinų kiekį spektrofotometriniu metodu. Nustatyti pasirinktų preparatų antioksidacinį aktyvumą DPPH metodu. Tyrimo objektai ir metodai: Preparatai su mėlynių uogų ekstraktu ir mėlynių uogų milteliais. Spektrofotometrinis metodas naudotas analizei: bendras fenolinių junginių kiekis – pagal Galo rūgšties kalibracinę kreivę; bendras antocianinų kiekis – pH diferenciniu metodu; antioksidacinis aktyvumas – DPPH laisvojo radikalo inaktyvavimo kreivė. Tyrimo rezultatai ir išvados: Iki 2013 m. birželio mėn. nustatyti 47 Lietuvoje registruoti mėlynių uogų preparatai. Į 97 proc. šių preparatų sudėtį įeinančių mėlynių uogų žaliavos forma – ekstraktas. 19 proc. visų tirtųjų sudaro kompleksiniai ir 9 proc. vienkomponenčiai preparatai. Spektrofotometriškai nustačius fenolinius junginius, didžiausias jų kiekis yra ,,Mėlynių ekstraktas“, kapsulėse (,,Vitaforce“, Jungtinė Karalystė), mažiausias - ,,Visioblue strong“, kapsulėse (,,Hankintatukku Oy“, Suomija). Preparatas ,,Mėlynių uogų ekstrakto tabletės su cinku forte 0,25 g”, („Elit-farm", Ukraina) eksperimentinio tyrimo metu nedavė apčiuopiamo rezultato... [toliau žr. visą tekstą] / Purpose of the work: To make the comparable ingredients and technology analysis of selected blueberry preparations registered in Lithuania, evaluate quantitatively and determine the antioxidant activity. Tasks: to investigate the supply of blueberry preparations registered in Lithuania. To make the comparable ingredients analysis of preparations mentioned above. Determine the phenolic compounds and anthocyanins quantity using the spectrophotometry method. Measure the antioxidant activity of selected preparation using DPPH method. Research objects and methodology: Preparations with blueberry extract and blueberry powder. Spectrophotometric methods were used for analysis: total phenolic content according calibration curve of galic acid; total anthocyanins - pH differential method; antioxidant activity - DPPH free radical inhibition. Results and conclusions: Until June, 2013 y. Lithuania set 47 record Blueberry fruit preparations. There was noticed that in 97 percent composition of the preparation, blueberry raw material shape - extract. 91 percent of explored preparations are multicomponent and just 9 percent - one-component. Determined by spectrophotometry, that the highest levels of the phenolic compounds are in ”Mėlynių ekstraktas“, capsules of (“Vitaforce”, United Kingdom) and the lowest – “Visioblue strong’’, capsules of (”Hankintatukku Oy’’, Finland). The quantity of Anthocyanins per recommended daily dose in the preparation was studied by spectrophotometry method. The... [to full text]

Tyrimo objektas ir metodai: šermukšnių (Sorbus L.) rūšių ir veislių lapų fenolinių junginių kiekybinės sudėties ir antioksidantinio aktyvumo tyrimas. Antioksidantinis aktyvumas nustatytas ABTS ir FRAP spektrofotometriniais bei efektyviosios skysčių chromatografijos pokolonėliniais metodais (ESC-ABTS ir ESC-FRAP). Darbo tikslas: ištirti šermukšnio (Sorbus L.) genties augalų lapų bendrą fenolinių junginių kiekį ir jo įvairavimą atskirose rūšyse ir veislėse bei įvertinti ekstraktų antioksidantinį aktyvumą spektrofotometriniais ir ESC pokolonėliniais metodais. Darbo uždaviniai: Nustatyti bendrą fenolinių junginių kiekį šermukšnių augalinėse žaliavose Folin-Ciocalteau metodu. Atlikti šermukšnio genties rūšių lapų kokybinę ir kiekybinę fenolinių junginių analizę ESC metodu. Įvertinti ekstraktų antiradikalinį aktyvumą ABTS spektrofotometriniu metodu. Atlikti ekstraktų redukcinių savybių tyrimą FRAP spektrofotometriniu metodu. Įvertinti atskirų junginių indėlį į bendrą ekstrakto antioksidantinį aktyvumą ESC-ABTS ir ESC-FRAP pokolonėliniais metodais. Įvertinti koreliacinį ryšį tarp bendro fenolinių junginių kiekio ir ekstraktų antioksidantinio aktyvumo. Išvados: Spektrofotometrinės analizės metu didžiausi fenolinių junginių kiekiai nustatyti S. caucasica Zinserl. rūšies (82,52±0,94 mg/g) ir ‘Alaya krupnaya’ veislės (131,76±7,04 mg/g) lapų pavyzdžiuose. Didžiausias redukcinis aktyvumas FRAP metodu (TE, µmol/g) nustatytas S. caucasica Zinserl. rūšies ir ‘Alaya krupnaya’ veislės... [toliau žr. visą tekstą] / Object and methods: quantitative analysis of phenolic compounds in rowan (Sorbus L.) species and hybrids and evaluation of antioxidant activity. Antioxidant activity was assessed by ABTS and FRAP spectrophotometric methods and by high performance liquid chromatography post-column reaction. Aim: to investigate quantitative composition of phenolic compounds and its variance in different rowan (Sorbus L.) species and hybrids and assess antioxidant activity of extracts by spectrophotometric and chromatographic methods. Objective: Evaluate quantitative composition of phenolic compounds in Sorbus L. species and hybrids leaves by Folin-Ciocalteau assay. Evaluate qualitative and quantitative composition of phenolic compounds in rowan species by high performance liquid chromatography. Assess free radical scavenging activity and determine reduction power of rowan leaves extracts by ABTS and FRAP spectrophotometric assay. Estimate correlation between quantity of phenolics and antioxidant activity in Sorbus L. leaves extracts. Assess input of individual compounds in overall antioxidant activity of extracts evaluated by HPLC-ABTS and HPLC-FRAP post-column methods. Results: In spectrophotometric analysis, the highest amount of phenolic compounds were determined in S.caucasica Zinserl. (82,52±0,94 mg/g) and ‘Alaya krupnaya’ (131,76±7,04 mg/g) leaves extracts. The highest antioxidant activity (TE, µmol/g) was observed in S.caucasica Zinserl. and ‘Alaya krupnaya’ ethanolic extracts using FRAP... [to full text]

[Truncated abstract. Please see the pdf format for the complete text.] Interest in the role of peroxynitrite in the pathogenesis of atherosclerosis has increased due to many in vitro studies which have demonstrated its potent oxidising and nitrating capability and immunohistochemical staining studies which demonstrate nitration of tyrosine in vivo. It is frequently suggested that the production of nitric oxide and superoxide at sites of inflammation implicates peroxynitrite as the major damaging reactive nitrogen species in vivo. Evidence for a role for peroxynitrite is often demonstrated by measurement of 3-nitrotyrosine yet even this cannot distinguish peroxynitrite from other nitrating species. Clearly, however, if peroxynitrite is important in atherogenesis, then identification of mechanisms for its detoxification could provide a means of preventing such effects. Therefore, this Thesis has sought to determine whether phenolic compounds of dietary origin can be preferentially nitrated by reactive nitrogen species thereby protecting endogenous structures, such as low density lipoproteins, from atherogenic modifications. This Thesis focuses upon phenolic acids as they have received relatively less attention than other classes of phenolic compounds, such as flavonoids, yet they are quite abundant in socially important beverages such as red wine. In order to complete the required analyses, the development of methods to detect phenolic acids and their nitration products together with 3-nitrotyrosine, dityrosine and 5-nitro-γ-tocopherol was necessary. The initial in vitro experiments described herein sought to determine the products of reaction of peroxynitrite with phenolic acids of the 4-hydroxy and 3,4-dihydroxy type and then to examine whether these products could account for a protective effect upon tyrosine, lipids and endogenous anti-oxidants, if any was observed, when isolated LDL was treated with SIN-1, which releases peroxynitrite through the simultaneous generation of nitric oxide and superoxide. A concurrent minor focus was to examine the relationship between structure and activity of these phenolic acids under various regimes of oxidative insult. These experiments indicate that, at least in this in vitro model, oxidation is a dominant mechanism over nitration. Peroxynitrite was shown to nitrate coumaric acid in moderate yields but exclusive oxidation of caffeic acid appeared to occur. Although a potential role for γ-tocopherol as an anti-nitration agent was inferred, all types of chemical treatment of LDL in the presence of phenolic acids yielded oxidation as the primary end point. In fact, nitration of tyrosine was not detected and nitration of coumaric acid was at the limit of detection. Since nitration of tyrosine is generally regarded as important in many disease states, a more physiological nitrating mechanism involving artificially stimulated neutrophils was used. This system demonstrated that although physiologically relevant reactive nitrogen species can result in nitration of phenolic compounds, in a complex system including biological structures (LDL) and phenolic compounds, oxidation but not nitration of all species appears to occur. As a consequence of the results above, an examination of carotid plaque was undertaken to determine to what extent nitration occurred relative to oxidation in atherosclerotic tissue. These studies applied methods developed herein to detect 3-nitrotyrosine and dityrosine in complex biological matrices as markers of nitration and oxidation respectively. The data obtained demonstrated that nitration was a minor modification of protein (0.01%) compared to oxidation (0.3%) even in a highly diseased tissue such as carotid artery plaque. A secondary study examining plasma revealed that dityrosine, which has been implicated in irreversible albumin aggregation in chronic renal failure and more recently in heart disease, is elevated in chronic renal failure subjects compared to well matched controls. A separate examination of plasma from healthy subjects revealed that in both the fasting and post prandial state 3-nitrotyrosine could not be detected and, in fact, interfering species could be problematic in the GC-MS analysis of 3-nitrotyrosine. The lack of nitration of any substrate observed in vitro using reactive nitrogen species generated in the aqueous phase, the relative lack of nitration of tyrosine in plaque proteins and the lipophilicity of nitric oxide, the precursor of all reactive nitrogen species, suggested that nitration could be more closely associated with lipid structures. The known ability of γ-tocopherol to form 5-nitro-γ-tocopherol was used to probe this concept. The 5-nitro-γ-tocopherol content of lipid extracts obtained from carotid artery plaques was very high (30%). This indicated that nitration is predominantly a lipid phase phenomenon and that nitrating species are present in much greater abundance than oxidising species in vivo.

Thesis (MSc Food Sc)--University of Stellenbosch, 2011. / ENGLISH ABSTRACT: The sensory characteristics and chemical composition of rooibos infusions were analysed to establish the extent of variation in sensory attributes and compositional parameters, and investigate whether correlations exist between the sensory characteristics and the phenolic composition. To capture as much potential variation as possible, 69 rooibos samples were collected throughout the 2009 harvesting season from different production areas. The samples were also representative of the different quality grades (A, B, C or D) which are based on the appearance of the leaves, and the colour and flavour of the infusion. Quantitative descriptive analysis was used to develop sensory profiles for each of the rooibos infusions. Considerable variation in the sensory characteristics was observed, highlighting the need for standardised terminology that can be used to describe what is often referred to as “characteristic” rooibos flavour. Aroma, flavour, taste, and mouthfeel descriptors were generated and assembled into a rooibos sensory wheel. The most recurring attributes were selected to compile a rooibos sensory lexicon which provides a definition and a reference standard for clarification of each term. A combination of “honey”, “woody” and “fynbos-floral” notes with a slightly sweet taste and subtle astringency may be regarded as “characteristic” rooibos attributes indicative of good quality tea. Pleasant “caramel” and “fruity-sweet” notes were also observed in a number of infusions, whereas negative descriptors, associated with poor quality rooibos, included “green” and “hay” flavours. Large variation in the composition of rooibos infusions was revealed through the quantification of soluble solids (SS), total polyphenols (TP), tannins and 14 monomeric phenolic compounds, as well as spectrophotometric colour measurements. High quality rooibos was associated with higher levels of SS, TP, tannins and phenolic compounds than low quality rooibos. Correlations between the compositional parameters and sensory attributes of the infusions indicated that several non-volatile compounds, including enolphenylpyruvic acid-2-glucoside, quercetin-3-glucoside, and iso-orientin, were associated with the characteristic sweet taste of rooibos, while bitterness was related to certain flavonoids such as luteolin, quercetin and aspalathin. The only compound significantly correlated to astringency was rutin, although it is likely that aspalathin and several other monomeric flavonoids also contribute to astringency. The tannin content was not associated with astringency possibly because of the limitations associated with the tannin quantification method. To determine whether compositional changes resulting from steam pasteurisation of rooibos leaves influence the sensory quality of rooibos infusions, differences in the phenolic composition and sensory attributes of infusions, prepared from unpasteurised and pasteurised rooibos, were analysed. Steam pasteurisation significantly reduced the SS, TP and aspalathin content of rooibos infusions, as well as the absorbance, especially at a wavelength of 450 nm. It also resulted in significant reductions in the intensities of most of the aroma and flavour attributes, especially the “green” notes associated with low quality rooibos. After steam pasteurisation the prominent “green” flavour of certain samples was frequently replaced by a “hay” flavour. The taste attributes, sweetness and bitterness, remained unchanged, whereas the astringency of rooibos infusions decreased significantly. / AFRIKAANSE OPSOMMING: Die sensoriese eienskappe en chemiese samestelling van rooibostee is geanaliseer ten einde die mate van variasie van die sensoriese sowel as die chemiese profiel te bepaal, asook om vas te stel of korrelasies bestaan tussen die sensoriese eienskappe en die fenoliese samestelling van rooibostee. Om soveel as moontlik potensiële variasie in te sluit, is 69 rooibos monsters tydens die 2009 oesseisoen van verskillende produksiegebiede versamel. Die monsters is ook verteenwoordigend van die verskillende kwaliteitgrade (A, B, C en D), wat op grond van die voorkoms van die teeblare, en die kleur en geur van die tee toegeken is. Kwantitatiewe beskrywende analise is gebruik om ‘n sensoriese profiel vir elke rooibostee op te stel. Aansienlike variasie in die sensoriese eienskappe is waargeneem. Dit het die behoefte aan gestandaardiseerde terminologie onderskryf, wat gebruik kan word om die term “karakteristieke” rooibos geur in meer detail te omskryf. Spesifieke terme vir aroma, geur, smaak en mondgevoel is geformuleer, en gebruik om ‘n sensoriese wiel vir rooibostee saam te stel. Terme wat die meeste voorgekom het, is gekies om in ‘n sensoriese leksikon vervat te word wat ‘n definisie en ‘n verwysingstandaard vir elke term voorstel. Die “karakteristieke” sensoriese eienskappe van goeie kwaliteit rooibostee kan soos volg gedefinieer word: ’n kombinasie van “heuning”, “houtagtige” en “fynbos-blom” geure met ‘n effense soet smaak en ‘n sagte vrankheid. Aangename “karamel” en “vrugtige-soet” geure is ook waargeneem, terwyl terme soos “groen” en “hooi” met swak kwaliteit rooibostee geassosieer is. Groot variasie in die samestelling van rooibostee is verkry deur kwantifisering van die inhoud van oplosbare vastestowwe (SS), totale polifenole (TP), tanniene, en 14 monomeriese fenoliese verbindings, asook spektrofotometriese kleurmetings. Hoë kwaliteit rooibostee is geassosieer met hoër vlakke van SS, TP, tanniene en fenoliese verbindings in vergelyking met lae kwaliteit rooibos. Korrelasies tussen laasgenoemde parameters en die sensoriese eienskappe het aangedui dat sekere nie-vlugtige verbindings, bv. fenielpirodruiwesuurglukosied, kwersetien-3-glukosied en iso-orientien, geassosieer is met die kenmerkende soet smaak van rooibos, terwyl ‘n bitter smaak geassosieer is met spesifieke flavonoïede soos luteolien, kwersetien and aspalatien. Die enigste verbinding wat betekenisvol gekorreleer het met vrankheid was rutien, maar dit is waarskynlik dat aspalatien en ander monomeriese flavonoïede ook bygedra het tot vrankheid. Die tannieninhoud het nie verband gehou met vrankheid nie, vermoedelik as gevolg van die beperkinge van die chemiese metode vir tannienbepaling. Om te bepaal of stoompasteurisasie van rooibos die sensoriese kwaliteit en chemiese samestelling van rooibostee beïnvloed, is die verskille in fenoliese samestelling en sensoriese eienskappe van die tee, berei van ongepasteuriseerde en gepasteuriseerde rooibos, ondersoek. Stoompasteurisasie het ‘n betekenisvolle verlaging teweegebring van die SS, TP en aspalatien inhoud, sowel as van die absorbansie van die tee, veral by ‘n golflengte van 450 nm. Dit het ook gelei tot ‘n statisties betekenisvolle vermindering van die intensiteit van die meeste aroma en geur eienskappe, veral die sogenaamde “groen” eienskap wat met swak kwaliteit rooibos verbind word. As gevolg van stoompasteurisasie is die opmerklike “groen” geur van sekere ongepasteuriseerde monsters herhaaldelik vervang deur ‘n “hooi” geur. Die smaakeienskappe, soet en bitter, was onveranderd, terwyl vrankheid statisties betekenisvol afgeneem het as gevolg van pasteurisasie.

Made available in DSpace on 2014-06-11T19:23:30Z (GMT). No. of bitstreams: 0 Previous issue date: 2010-07-22Bitstream added on 2014-06-13T20:10:55Z : No. of bitstreams: 1 corniani_n_me_botfca.pdf: 1146521 bytes, checksum: fa6a3da1e11828289d5c9cb1e7208258 (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / As espécies do gênero Eucalyptus apresentam notável capacidade de extração de água e nutrientes, assimilação fotossintética e crescimento, permitindo seu cultivo em ambientes considerados impróprios para muitas outras espécies. Além dos fatores ambientais atuando adversamente no desempenho das plantas, atualmente estas estão sujeitas aos efeitos das mudanças ambientais globais causadas pelas atividades antrópicas, tais como o aumento na incidência da radiação ultravioleta-B (RUV-B) em conseqüência da destruição da camada de ozônio. Estudos recentes têm mostrado que a RUV-B promove aumento no nível de espécies reativas de oxigênio (ERO), ocasionando estresse oxidativo nas plantas. Entretanto, as plantas apresentam um sistema de defesa contra o estresse oxidativo, constituído de enzimas antioxidantes, tais como superóxido dismutase (SOD, EC 1.15.1.1) e peroxidase (POD, EC 1.11.1.7), além de outros compostos, como antocianinas, carotenóides e compostos fenólicos, que impedem a formação de ERO ou eliminam as já existentes. Há algum tempo, tem sido verificada em plantas a presença de óxido nítrico (NO), radical livre endógeno que possui a capacidade de controlar o nível e a toxicidade das ERO. Portanto, o objetivo deste trabalho foi investigar o possível efeito citoprotetor do NO em plantas de eucalipto (Eucalyptus urograndis) expostas à RUV-B. Assim, foi realizado um primeiro experimento no qual plantas de eucalipto foram expostas à RUV-B em baixa (controle) e elevada incidência, com o intuito de verificar se a radiação promove estresse oxidativo nesta espécie. Foram realizadas coletas de lâminas foliares aos cinco, dez e quinze dias após o início da exposição à RUV-B para posterior determinação do nível de peroxidação lipídica. A exposição por 15 dias à RUV-B... / The species of the genus Eucalyptus present notable capacity to extract water and nutrients, photosynthetic assimilation and growth, allowing cultivation in ambient considered inappropriate for many other species. Besides the environmental factors acting adversely in the performance of the plants, nowadays they are subject to the effects of the global environmental changes caused by human activities, such as the increase in the incidence of the ultraviolet-B radiation (UV-B) in consequence of the ozone layer destruction. Recent studies show that UV- B radiation promotes increase in the level of reactive oxygen species (ROS), causing oxidative stress in plants. However, plants have a defence system against oxidative stress, constituted of antioxidant enzymes, such as, superoxide dismutase (SOD, EC 1.15.1.1), peroxidase (POD, EC 1.11.1.7), and others compounds, such as anthocyanins, carotenoids and phenolic compounds, that avoid ROS formation or eliminate the already existent. There has been long, it was noticed the presence of nitric oxide (NO) in plants, endogenous free radical able to control the ROS level and toxicity. Therefore, the aim of this work was to investigate the possible citoprotector effect of the NO in eucalyptus (Eucalyptus urograndis) plants subjected to UV-B radiation. So, a first experiment was setup in which plants of eucalyptus were subjected to radiation in low (control) and high incidence of UV-B, to verify if UV-B radiation promotes oxidative stress in this specie. Leaves were collected at five, ten and fifteen days after the beginning of exposure to UV-B for subsequent evaluation of the level of lipid peroxidation. Exposure for 15 days to RUV-B increased the lipoperoxide content in the eucalyptus... (Complete abstract click electronic access below)

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / In the recent years, the concern about wines with better sensory characteristics and biological properties has motivated many studies on the phenolic compounds. The knowledge of ripening process of these substances is related to the necessity to produce high quality wines. In this study, red grape varieties VitisviniferaMalbec, Shiraz and Tannat were evaluated in relation to their phenolic ripeness. In the first two varieties, three harvest dates of a same vineyard were evaluated. In Tannat variety, one harvest date for three different vineyards was compared. All vineyards are situated in Rio Grande do Sul, Brazil, and they were studied in the crops of 2009/2010 and 2010/2011. The grape berries were evaluated in relation to the rate of skins, seeds, and pulps; and in relation to the industrial (pH, Brix and total acidity) and phenolic ripeness. For this last evaluation, 125 berries were crushed and macerated at pH 1.0 and pH 3.2 for 4 hours. Then, the following parameters were evaluated: extractability of anthocyanins (EA%), tannins in the skins (dpell%), tannins in the seeds (Mp%), total polyphenolic index (TPI 280), total anthocyanins (ApH1,0) and extractable anthocyanins (ApH3,2). Malbec and Shiraz varieties were micro-vinified by traditional method of red vinification, then the final wine was evaluated for its physic-chemical properties (pH and total acidity) and the content of phenolic compounds (IPT, total polyphenols, tannins, and anthocyanins). The analyses were performed in triplicate and statistically evaluated by the Tukey test at 5% of probability, by Principal Component Analysis (PCA) and by linear correlation. For Malbec variety, the outcomes showed a bigger industrial and phenolic ripeness (more Brix, less Total Acidity, more IPT, more content of anthocyanins and low values of Mp% and dpell%) to the 2010/2011 crop and later harvests. The Shiraz variety showed higher levels of total anthocyanins, extractable anthocyanins, and IPT in the second harvest date. The phenolic ripeness in the second crop was also higher for this variety, which had a higher extractability of anthocyanins (less EA%) and a great ripeness of tannins from the seeds and skins (lower Mp% and higher dpell%, respectively). For Tannat variety, the sample of Itaqui (warmer region) showed a higher content of anthocyanins and IPT in the first crop, which is similar to the sample of Dom Pedrito (less warmer region than Itaqui) in the following crop. In PCA, the samples were amounted by the influence of crop and the harvest date. The wine of both varieties presented higher levels of total phenolic, anthocyanins, and IPT in the vinification of later harvest and the crop of 2010/2011. The total polyphenol content, IPT 280, and the content of anthocyanins in wines were highly correlated with the rate of the industrial ripeness of the grape, content of extractable anthocyanins (ApH 3.2), tannin content in skins (Mp%). It is possible to conclude that later harvest can provide a grape with a higher phenolic ripeness. However, this ripeness is closely related and dependent on appropriate weather conditions of harvest. / Nos últimos anos, a preocupação por vinhos com melhores características sensoriais e propriedades biológicas tem motivado inúmeros estudos sobre os compostos fenólicos. O conhecimento sobre o processo de maturação dessas substâncias vem ao encontro da necessidade de se produzir um vinho de maior qualidade. Neste estudo, as variedades tintas Vitis vinifera Malbec, Shiraz e Tannat foram avaliadas quanto à sua maturação fenólica. Para as duas primeiras variedades, avaliaram-se três datas de colheita de um mesmo vinhedo. Para a variedade Tannat, comparou-se uma data de colheita de três vinhedos distintos. Todos os vinhedos localizam-se no Estado do Rio Grande do Sul e foram estudados nas safras de 2009-2010 e 2010-2011. As bagas de uva foram avaliadas quanto à proporção de cascas, sementes e polpa, quanto à maturação industrial (pH, Brix e Acidez Total) e quanto à maturação fenólica. Para esta última avaliação, 125 bagas foram triturados e macerados em pH 1,0 e pH 3,2 por 4 horas; após, avaliou-se os seguintes parâmetros: extratibilidade de antocianinas (EA%), taninos nas cascas (dpell%), taninos nas sementes (Mp%), Índice Polifenólico Total (IPT 280), antocianinas totais (ApH1,0) e antocianinas extraíveis (ApH3,2). As variedades Malbec e Shiraz foram microvinificadas pelo método tradicional de vinificação em tinto, e o vinho final foi avaliado quanto às propriedades físico-químicas (pH e Acidez Total) e ao teor de compostos fenólicos (IPT280, Polifenóis Totais, Taninos e Antocianinas). As análises foram realizadas em triplicata e estatisticamente avaliadas pelo teste de Tukey a 5% de probabilidade, por Análise de Componente Principal (PCA) e por correlação linear. Para a variedade Malbec, os resultados evidenciaram maior maturação industrial e fenólica (maior Brix, menor Acidez Total, maior IPT280, maior teor de antocianinas e dpell%e baixos valores de Mp%) para a safra de 2010-2011 e para as colheitas mais tardias. A variedade Shiraz demonstrou maiores teores de antocianinas totais, antocianinas extraíveis e IPT na segunda data de colheita. A maturação fenólica na segunda safra também foi superior para esta variedade, observando-se maior extratibilidade de antocianinas (menor EA%) e maior maturação dos taninos das sementes e das cascas (menor Mp% e maior dpell%, respectivamente). Para a variedade Tannat, a amostra de Itaqui (região de clima mais quente) apresentou maior teor de antocianinas e IPT280 na primeira safra, equiparando-se à amostra de Dom Pedrito (região de clima menos quente que Itaqui) na safra seguinte. Na PCA, as amostras foram agrupadas pela influência da safra e data de colheita. Os vinhos de ambas as variedades apresentaram maiores teores de compostos fenólicos totais, antocianinas e IPT280 nas vinificações de colheitas mais tardias e na safra de 2010-2011. O conteúdo de polifenóis totais, IPT 280 e o conteúdo de antocianinas nos vinhos foram altamente correlacionados com os índices de maturação industrial da uva, teor de antocianinas extraíveis (ApH 3,2), teor de taninos nas cascas (dpell%) e teor de taninos nas sementes (Mp%). Conclui-se que colheitas mais tardias podem proporcionar uma uva de maior maturação fenólica. Porém, esta maturação está intimamente relacionada e dependente de adequadas condições climáticas da safra.

O pinhão é a semente comestível da Araucaria angustifolia, sendo consumido no sul e sudeste do Brasil. Possui excelentes características nutritivas e energéticas. As sementes têm uma atividade de água muito elevada (>0,98), o que facilita o desenvolvimento de fungos durante a estocagem, dificultando a sua comercialização. A literatura sobre os seus aspectos nutricionais e tecnológicos é muito escassa. O objetivo principal deste trabalho foi determinar aspectos tecnológicos e de engenharia relacionados com o processamento e armazenamento do pinhão. Foi estudada a vida de prateleira do pinhão cru a 25°C (5 dias), 5°C (3 meses) e –18°C (mais de 8 meses) através de análises fisicoquímicas, microbiológicas e sensoriais. Foram propostas as seguintes operações preliminares para preservar a qualidade da semente comercializada: recepção, limpeza, seleção, classificação e armazenamento. Foram determinadas as isotermas de desorção do pinhão cru e cozido e de adsorção da farinha de pinhão cru em diferentes temperaturas (entre 10 e 40°C) sendo ajustados modelos da literatura e calculadas a entalpia e entropia diferenciais. Para os três casos o modelo que melhor se ajustou aos dados foi o de Chirife e a teoria compensatória foi aplicada. Foi avaliada também a cinética de secagem do pinhão cru e cozido em secador de bandeja a 55, 70 e 85°C. Os dados foram ajustados a modelos de secagem e calculadas as difusividades efetivas, sendo que a difusividade do pinhão cozido (1,18 a 3,17 x10-10 m2s-1) foi menor do que a do pinhão cru (1,64 a 4,52 x10-10 m2s-1), provavelmente devido à gelatinização do amido. Um dos processos empregados pela tecnologia de alimentos, com a finalidade de obter produtos estáveis a partir de sementes, é a produção de farinhas. Neste trabalho foi produzida farinha de pinhão variando as seguintes condições de processo: utilização de pinhão cru e cozido e diferentes temperaturas de secagem (55, 70 e 85°C). As farinhas produzidas com pinhão cozido apresentaram cor mais escura, com uma luminosidade menor (Lmédia= 75.8) e coordenada de cromaticidade maior (amédia= 6.9), quando comparadas com as farinhas de pinhão cru (Lmédia= 89.0, amédia= 2.1). A temperatura de secagem também influenciou a cor das farinhas produzidas com pinhão cru. Através de análise multivariada dos dados foi possível agrupar as amostras. A partir dos resultados da análise sensorial constatou-se que o suflê produzido com a farinha de pinhão cozido apresentou maior aceitação, obtendo-se um produto que une praticidade, inovação e apelo regional. Por último, como uma forma de aproveitamento da casca do pinhão, foi estudada a extração de compostos fenólicos da casca, como uma nova fonte de antioxidantes. Foram determinadas as melhores condições em termos de volume e concentração de solvente (etanol) e temperatura de extração, utilizando a metodologia de superfície de resposta. / The seeds of Araucaria angustifolia, named pinhão, are consumed in the South and Southeast of Brazil. Pinhão has good nutritious and energetic characteristics. Since the seeds have a high water activity (>0,98), they can be easily contaminated by mushrooms during the stockpiling, hindering its commercialisation. The literature about nutritional and technological aspects of pinhão is very scarce. The main objective of this work was to determine technological and engineering aspects of the processing and storage of pinhão. The Shelf life of raw pinhão was studied at 25°C (5 days), 5°C (3 months) and -18°C (more than 8 months) using physiochemical, microbiological and sensorial analyses. The following preliminary operations were proposed to preserve the quality of the seed for commercialisation: reception, cleaning, selection, classification and storage. Desorption of raw and cooked pinhão and adsorption isotherms of raw pinhão flour were determined at different temperatures (between 10 and 40°C), modelled using well know isotherm models and differential enthalpy and entropy were calculated. Results show that the Chirife model most appropriately represents the experimental data for the three cases and the enthalpy-entropy compensation theory was applied. Drying behavior of raw and cooked pinhão was studied at 55, 70 and 85°C. The data were adjusted to drying models and effective diffusivity were calculated. Values for cooked pinhão (1,18 to 3,17 x10-10 m2s-1) were smaller than values for raw pinhão (1,64 to 4,52 x10-10 m2s-1), probably due to starch gelatinisation. One of the processes used by food technology with the purpose of obtaining stable products from seeds, it is the production of flours. In this work pinhão flour was produced varying the following process conditions: use of raw or cooked pinhão and different drying temperatures (55, 70 and 85°C). Flours produced with cooked pinhão presented darker colour, with a smaller brightness (Lmean = 75.8) and larger coordinate of chromaticity (amean = 6.9), when compared with raw pinhão flours (Lmean = 89.0, amean = 2.1). Drying temperature also influenced the colour of flours produced with raw pinhão. Through multivariate analysis of the data it was possible to cluster the samples. Results of sensorial analysis show that soufflé produced with cooked pinhão flour presented larger acceptance, being obtained a new product with regional appeal. The extraction of phenolic compounds of pinhão coats was also studied as a new source of antioxidants. The best condition of solvent volume, ethanol concentration and temperature, using factorial design and response surface methodology, were obtained.

Submitted by ANA PAULA MAIA PAIVA (anapaula.agro@ymail.com) on 2018-10-18T14:38:52Z No. of bitstreams: 1 TESE ANA PAULA MAIA PAIVA-Final.pdf: 2467388 bytes, checksum: c5d2a5d139387fb46d6c4ecbefc34c9d (MD5) / Approved for entry into archive by Ana Lucia de Grava Kempinas (algkempinas@fca.unesp.br) on 2018-10-18T17:16:16Z (GMT) No. of bitstreams: 1 paiva_apm_dr_botfca.pdf: 2471505 bytes, checksum: e8cdc3ef558b842f8b02e93e3f5dd2f6 (MD5) / Made available in DSpace on 2018-10-18T17:16:16Z (GMT). No. of bitstreams: 1 paiva_apm_dr_botfca.pdf: 2471505 bytes, checksum: e8cdc3ef558b842f8b02e93e3f5dd2f6 (MD5) Previous issue date: 2018-01-03 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / O objetivo desse trabalho foi avaliar cultivares de uva para suco e vinho, cultivadas em São Manuel, SP, nas variáveis de fenologia, evolução da maturação, produção, características físicas, químicas e bioquímicas nas bagas e cachos. O experimento foi conduzido na Fazenda Experimental de Ensino, Pesquisa e Produção de São Manuel, SP, pertencente à Faculdade de Ciências Agronômicas, Campus de Botucatu FCA/UNESP, no Laboratório de Fruticultura do Departamento de Horticultura da FCA/UNESP e no Laboratório de Química e Bioquímica Vegetal do Departamento de Química e Bioquímica do Instituto de Biociências do IBB/UNESP, em Botucatu-SP, no período de agosto de 2014 a janeiro de 2017. Os tratamentos consistiram em quatro cultivares de uvas para suco e vinho (BRS Violeta, Bordô, BRS Cora e BRS Carmem) sobre o porta-enxerto ‘IAC 572’. Foram avaliadas a produção e produtividade; massas frescas de cacho (MFC) e engaço (MFE); comprimento e largura de cachos e engaços; número de bagas por cacho; massa fresca (MFB), comprimento (CB) e largura de bagas (LB); número e massa fresca de sementes por baga (MFSB); relação MFSB/MFB; sólidos solúveis (SS), acidez titulável (AT), relação SS/AT; pH, antocianinas monoméricas totais, polifenóis totais, flavonoides totais e atividade antioxidante das uvas. Os dados obtidos foram submetidos à análise de variância e as médias comparadas pelo teste Tukey, a 5% de probabilidade. Embora com variações entre os ciclos produtivos, as cultivares BRS Violeta e BRS Cora apresentaram maiores produção e produtividade nos ciclos avaliados. Por sua vez, apesar das boas características físicas de cachos da cultivar BRS Carmem, essa videira juntamente com as videiras da cultivar Bordô apresentaram baixo potencial produtivo, o que pode ser um fator limitante para seu cultivo na região de estudo. Apesar dos altos níveis de acidez titulável em todas as uvas produzidas no segundo ciclo, pode-se afirmar que as condições climáticas na região de São Manuel proporcionaram boas características químicas das uvas, que apresentaram teor de sólidos solúveis mínimos para a elaboração de sucos. Ainda assim a uva ‘BRS Cora’ manteve altos níveis de acidez titulável. A avaliação do cultivo das diferentes cultivares para a elaboração de suco e/ou vinho de mesa, mostrou ter potencial, pois os possíveis desequilíbrios de cada uma podem ser corrigidos por meio de cortes durante o processamento. Uvas da cultivar BRS Violeta apresentaram os maiores teores de polifenóis totais e maior atividade antioxidante. Alto conteúdo desses compostos também foi encontrado nas uvas Bordô e BRS Cora. O conteúdo de antocianinas monoméricas totais, polifenóis totais e, principalmente, de flavonoides totais está correlacionada positivamente à atividade antioxidante das uvas. / The objective of this work was to evaluate grape cultivars for juice and wine, cultivated in São Manuel, SP, in phenology, production, physical, chemical and biochemical characteristics. The experiment was carried out at the Experimental Farm for Teaching Research and Production of São Manuel, SP, belonging to the Faculty of Agronomic Sciences, Campus de Botucatu FCA / UNESP, at the Fruit Laboratory of the Department of Horticulture of FCA / UNESP and at the Laboratory of Chemistry and Plant Biochemistry of the Department of Chemistry and Biochemistry of the Institute of Biosciences of IBB / UNESP, in Botucatu-SP, from August 2014 to January 2017. The treatments consisted of four cultivars of grapes for juice and wine (BRS Violeta, Bordô, BRS Cora and BRS Carmem) on the rootstock 'IAC 572'. Production and productivity were evaluated; Fresh bunch (MFC) and stalks (MFE); Length and width of bunches and stalks; Number of berries per cluster; Fresh mass (MFB), length (CB) and width of berries (LB); Number and fresh mass of seeds per berry (MFSB); MFSB / MFB ratio; Soluble solids (SS), titratable acidity (AT), SS / AT ratio; PH, total monomeric anthocyanins, total polyphenols, total flavonoids and antioxidant activity of grapes. The data were submitted to analysis of variance and the means were compared by the Tukey test, at 5% probability. Although with variations between the productive cycles, cultivars BRS Violeta and BRS Cora presented higher production and productivity in the evaluated cycles. On the other hand, in spite of the good physical characteristics of curls of the BRS Carmem cultivar, this vine along with the vines of the cultivar Bordô presented / displayed low productive potential, what can be a limiting factor for its cultivation in the region of study. In spite of the high levels of titratable acidity in all the grapes produced in the second cycle, it can be stated that the climatic conditions in the São Manuel region provided good chemical characteristics of the grapes, which presented minimum soluble solids content for juice production. Still the 'BRS Cora' grape maintained high levels of titratable acidity. The evaluation of the cultivation of the different cultivars for the elaboration of juice and / or table wine, showed to have potential, because the possible imbalances of each one can be corrected through cuts during the processing. Grapes of the cultivar BRS Violeta showed the highest total polyphenol content and higher antioxidant activity. High content of these compounds was also found in Bordô and BRS Cora grapes. The content of total monomeric anthocyanins, total polyphenols and, mainly, total flavonoids correlates mainly with the antioxidant activity of grapes. / 001