Academic literature on the topic 'Flavour ester'

Lipases are one of the most important industrial biocatalyst which catalyzes the hydrolysis of lipids. It can also reverse the reaction at minimum water activity. Because of this pliable nature, it is widely exploited to catalyze the diverse bioconversion reactions, such as hydrolysis, esterification, interesterification, alcoholysis, acidolysis and aminolysis. The property to synthesize the esters from the fatty acids and glycerol promotes its use in various ester synthesis. The esters synthesized by lipase finds applications in numerous fields such as biodiesel production, resolution of the recemic drugs, fat and lipid modification, flavour synthesis, synthesis of enantiopure pharmaceuticals and nutraceuticals. It plays a crucial role in the food processing industries since the process is unaffected by the unwanted side products. Lipase modifications such as the surfactant coating, molecular imprinting to suit for the non-aqueous ester synthesis have also been reported. This review deals with lipase catalyzed ester synthesis, esterification strategies, optimum conditions and their applications in food processing industries.

In the course of the present study, filter layers with embedded zeolites (patented layer of the company Filtrox (Zwingen, Switzerland) with the brand name Fibrafix® TX) were tested for the elimination of the wine defects, cork taint (2,4,6-trichloroanisole), mouldy aroma (geosmin), and strong “animal” phenolic aromas (4-ethylphenol, 4-ethylguaiacol, 4-ethylcatechol). The test design allowed a comparison with conventional filter layers (sterile filter layers) in a small-scale trial (25 L) as well as in a large-scale trial (125 L). By means of gas chromatography-mass spectroscopy, not only the impact compounds of the wine faults were analysed, but also the loss of volatile substances such as 15 free monoterpenes, 34 ester compounds, and 24 wood flavours. Sensory analyses were carried out by means of expert panels. The Fibrafix® TX layers were satisfactory with regard to the reduction in 2,4,6-trichloroanisole (the lead substance of cork flavour; reduction > 90%) and geosmin (the lead substance of mould flavour; reduction > 75%), but not with regard to the reduction in 4-ethylphenol, 4-ethylguaicol, and 4-ethylcatechol. However, the reduction in these off-flavours was generally not specific enough to prevent a loss of volatile compounds, especially various ester compounds and free monoterpenes. The wines obtained after filtration (in case of contamination with geosmin or TCA) were sensory faultless, but for some of the tasters, the overall quality of these wines did not correspond to the quality wine clasification.

In the course of the present study, filter layers with embedded zeolites (patented layer of the company Filtrox (Zwingen, Switzerland) with the brand name Fibrafix® TX) were tested for the elimination of the wine defects, cork taint (2,4,6-trichloroanisole), mouldy aroma (geosmin), and strong “animal” phenolic aromas (4-ethylphenol, 4-ethylguaiacol, 4-ethylcatechol). The test design allowed a comparison with conventional filter layers (sterile filter layers) in a small-scale trial (25 L) as well as in a large-scale trial (125 L). By means of gas chromatography-mass spectroscopy, not only the impact compounds of the wine faults were analysed, but also the loss of volatile substances such as 15 free monoterpenes, 34 ester compounds, and 24 wood flavours. Sensory analyses were carried out by means of expert panels. The Fibrafix® TX layers were satisfactory with regard to the reduction in 2,4,6-trichloroanisole (the lead substance of cork flavour; reduction > 90%) and geosmin (the lead substance of mould flavour; reduction > 75%), but not with regard to the reduction in 4-ethylphenol, 4-ethylguaicol, and 4-ethylcatechol. However, the reduction in these off-flavours was generally not specific enough to prevent a loss of volatile compounds, especially various ester compounds and free monoterpenes. The wines obtained after filtration (in case of contamination with geosmin or TCA) were sensory faultless, but for some of the tasters, the overall quality of these wines did not correspond to the quality wine clasification.

Abstract Button mushroom (Agaricus bisporus) is sold well for its unique flavour and nutritional benefits. However, the mushroom flavour deteriorates quickly during storage because of its delicate structure and high moisture. In this study, the effects of nitric oxide (NO) application on flavour compounds and antioxidant enzyme activities of stored button mushrooms were investigated. The button mushrooms were immersed in the NO donor sodium nitroprusside (15 μmol/L) for 3 min and then stored under the condition of 4 °C, 90% relative humidity for 12 days. Results showed that the treated mushrooms have reduced weight loss rate, uniform white colour, and higher firmness during storage. Compared to the control, the ketones, alcohols, esters, and aldehydes in the NO-treated button mushroom increased sharply at 3 days of storage and then showed a continuing decline trend, except ester compounds which reached the peak value at 6 days of storage. In addition, NO treatment increased the total phenolics and catalase activity and inhibited the polyphenol oxidase activity in the stored button mushroom. These results indicated that NO treatment is an alternative storage technology to enhance antioxidant capacity and maintain flavour and consumer acceptance of stored button mushroom.

Medium chain esters produced from fruits and flowering plants have a number of commercial applications including use as flavour and fragrance ingredients, biofuels, and in pharmaceutical formulations. These esters are typically made via the activity of an alcohol acyl transferase (AAT) enzyme which catalyses the condensation of an alcohol and an acyl-CoA. Developing a microbial platform for medium chain ester production using AAT activity presents several obstacles, including the low product specificity of these enzymes for the desired ester and/or low endogenous substrate availability. In this study, we engineeredEscherichia colifor the production of butyl octanoate from endogenously produced octanoyl-CoA. This was achieved through rational protein engineering of an AAT enzyme fromActinidia chinensisfor improved octanoyl-CoA substrate specificity and metabolic engineering ofE. colifatty acid metabolism for increased endogenous octanoyl-CoA availability. This resulted in accumulation of 3.3 + 0.1 mg/L butyl octanoate as the sole product fromE. coliafter 48 h. This study represents a preliminary examination of the feasibility of developingE. coliplatforms for the synthesis single medium chain esters from endogenous fatty acids.

Traditionally, ugba of acceptable quality made from African oil bean (Pentaclethra macrophylla Benth) is characterized by its typical flavour and aroma. In this study, aroma profile and volatile flavour compounds released by spontaneously fermented ugba and five samples of ugba inoculated with starter cultures (A - Bacillus subtilis; B - B. safensis; C- Mixed culture of B. subtilis and B. safensis; D - Mixed culture of B. subtilis, B. safensis, B. clausii, B. licheniformis, and B. xylanilyticus; and E - B. cereus sensu lato) were monitored at 24 hour interval during 72 hours fermentation using solid-phase microextraction (SPME) and gas chromatography-mass spectrometry (GC-MS). Commercially prepared ugba is the control. The total heterotrophic bacterial count (THBC) of the samples were determined using Standard Microbiological Methods; the pH involved the use of standard method. Increase in the THBC and pH of all the samples of fermenting ugba were within the range of 1.4 × 104 to 1.7 x 1012 CFU/ml and 5.89 to 8.24, respectively. A total of 31 compounds which comprise of 7 ketones, 7 alcohols, 5 organic acids, 4 aldehydes, 2 pyrazines, 1 ester, 1 furan, 1 sulfur and 3 other compounds were identified as aroma compounds in the fermenting samples of ugba. Flavour compounds categorized as ester, furan and sulfur compound were consistently detected in all the samples of ugba prepared in the laboratory. Ugba prepared using starter cultures including the spontaneously fermented sample compare favourably with commercially produced ugba with regards to volatile flavour compounds.

This study compared the use of three sensory and analytical techniques: Quantitative Descriptive Analysis (QDA), Napping, and Gas Chromatography-Mass Spectrometry (GC-MS) for the assessment of flavour in nine unmatured whisky spirits produced using different yeasts. Hierarchical Multiple Factor Analysis (HMFA) showed a similar pattern of sample discrimination (RV scores: 0.895–0.927) across the techniques: spirits were mostly separated by their Alcohol by Volume (ABV). Low ABV spirits tended to have heavier flavour characteristics (feinty, cereal, sour, oily, sulphury) than high ABV spirits, which were lighter in character (fruity, sweet, floral, solventy, soapy). QDA differentiated best between low ABV spirits and GC-MS between high ABV spirits, with Napping having the lowest resolution. QDA was time-consuming but provided quantitative flavour profiles of each spirit that could be readily compared. Napping, although quicker, gave an overview of the flavour differences of the spirits, while GC-MS provided semi-quantitative ratios of 96 flavour compounds for differentiating between spirits. Ester, arenes and certain alcohols were found in higher concentrations in high ABV spirits and other alcohols and aldehydes in low ABV spirits. The most comprehensive insights on spirit flavour differences produced by different yeast strains are obtained through the application of a combination of approaches.

Flavour in Scotch malt whisky is a key differentiating factor for consumers and producers alike. Yeast (commonly Saccharomyces cerevisiae) metabolites produce a significant amount of this flavour as part of distillery fermentations, as well as ethanol and carbon dioxide. Whilst yeast strains contribute flavour, there is limited information on the relationship between yeast strain and observed flavour profile. In this work, the impact of yeast strain on the aroma profile of new make spirit (freshly distilled, unmatured spirit) was investigated using 24 commercially available active dried yeast strains. The contribution of alcoholic, fruity, sulfury and sweet notes to new make spirit by yeast was confirmed. Generally, distilling strains could be distinguished from brewing and wine strains based on aroma and ester concentrations. However, no statistically significant differences between individual yeast strains could be perceived in the intensity of seven aroma categories typically associated with whisky. Overall, from the yeast strains assessed, it was found that new make spirit produced using yeast strains marketed as ‘brewing’ strains was preferred in terms of acceptability rating.

Tofu from Lembang, Tofu from Sumedang and Tofu from Garut are three types tofu that are produced in different places, but still close. Although made from the same raw material that is soy, every tofu will produce a different tofu, both in terms of taste, aroma, texture and suppleness. Differences in the type of flavour compouds used can also be a factor causing divergence of any tofu product. The characteristic of the tofu were determined by identifying the flavour compouds GC-MS, as well as organoleptic tests. To determine differences in compound-producing falvour in the tofu. The results extraction method was carried out on tofu samples using n-pentane as solvent then analyzed using GC-MS. Flavour compounds of all three types of tofu included into the ester group, alkenes and aldehydes. Organoleptic tests show that the panelists prefered. Tofu from Sumedang for test of colour appearance (82%), for the test texture panelists like Tofu from Garut (83%), for test of elasticity panelists prefered Tofu from Garut Lembang (75%), for taste test panelists prefered Tofu from Lembang (88%), panelists prefered Tofu from Lembang for flavour test (82%).

Gli esteri svolgono un ruolo rilevante nell'industria alimentare, sono tra i composti più importanti e versatili di aromi e fragranze naturali in alimenti, bevande e cosmetici. La loro preparazione da substrati naturali e l'utilizzo di bioprocessi (eg. fermentazione o reazioni enzimatiche) è attrattivo, perché il prodotto finale può essere etichettato e commercializzato in UE e USA come naturale. Pertanto, nuovi approcci biotecnologici per ottenere aromi sono desiderati, una volta che siano efficienti e sostenibili. Molti esteri con proprietà aromatiche possono essere ottenuti enzimaticamente usando lipasi che catalizzano reazioni di esterificazione, transesterificazione o interesterificazione. In questa tesi di dottorato abbiamo sviluppato due sistemi per la produzione di esteri con proprietà aromatiche: 1) Un metodo biocatalitico per la preparazione enzimatica di diversi esteri con proprietà aromatiche da alcoli primari (isoamilico, n-esilico, geranilico, cinnamilico, 2-feniletilico e benzilico) ed esteri etilici naturalmente disponibili (formiato, acetato, propionato e butirrato). Le biotrasformazioni sono catalizzate da un'aciltransferasi di Mycobacterium smegmatis (MsAcT) e precedute da eccellenti rese (80- 97%) e brevi tempi di reazione (30-120 minuti), anche quando sono state utilizzate concentrazioni di substrato elevate (fino a 0,5 M). Questa strategia enzimatica rappresenta un'alternativa efficace all'applicazione delle lipasi nei solventi organici e un miglioramento significativo rispetto ai metodi già noti in termini di uso ridotto di solventi organici, aprendo la strada a una preparazione sostenibile ed efficiente degli agenti aromatizzanti naturali. 2) Un metodo biocatalitico con la lipasi legata al micelio liofilizzato di Aspergillus oryzae che catalizza l'esterificazione diretta di alcoli e acido acetico in solvente organico. Ha mostrato un'elevata stabilità verso substrati e prodotti. L'acqua prodotta durante l'esterificazione non ha influenzato in modo significativo l'equilibrio della reazione, consentendo conversioni elevate. Queste caratteristiche sono state sfruttate per preparare esteri con proprietà aromatiche dell'acetato (isoamil e cinnamil acetato) in sistemi in batch e continui. È stato sviluppato un continuous stirred tank membrane reactor (CST-MR) ovvero un reattore continuo a membrana sotto agitazione per garantire una buona produttività e un'elevata stabilità del biocatalizzatore. Entrambi i sistemi di produzione sono promettenti, rappresentano due diverse alternative e possono essere ulteriormente ottimizzati e scalati per gli interessi del settore.
Esters play a significant role in the food industry, they are among the most important and versatile components of natural flavours and fragrances in food, drinks and cosmetics Their preparation starting from natural substrates and using bioprocesses (e.g., fermentation or enzymatic reactions) is appealing, since the final product can be labelled and commercialized in EU and USA as natural. Therefore, new biotechnological approaches for obtaining flavours are highly demanded as long as they are efficient and sustainable. Many flavour/fragrance esters can be enzymatically obtained using lipases that catalyse esterification, transesterification or interesterification reactions. In this PhD thesis we studied two systems for production of flavours esters: 1) A straightforward biocatalytic method for the enzymatic preparation of different flavour esters starting from primary alcohols (e.g., isoamyl, n-hexyl, geranyl, cinnamyl, 2-phenethyl, and benzyl alcohols) and naturally available ethyl esters (e.g., formate, acetate, propionate, and butyrate) was developed. The biotransformations are catalysed by an acyltransferase from Mycobacterium smegmatis (MsAcT) and preceded with excellent yields (80-97%) and short reaction times (30-120 minutes), even when high substrate concentrations (up to 0.5 M) were used. This enzymatic strategy represents an efficient alternative to the application of lipases in organic solvents and a significant improvement compared to already known methods in terms of reduced use of organic solvents, paving the way to a sustainable and efficient preparation of natural flavouring agents. 2) Mycelium-bound lipase of dry mycelium of Aspergillus oryzae catalysed direct esterification of alcohols and acetic acid in organic solvent, showing high stability towards substrates and products. Water produced during the esterification did not significantly affect the equilibrium of the reaction, allowing for high conversions. These features were exploited for preparing flavour-active acetate esters (e.g., isoamyl and cinnamyl acetate) in batch and continuous systems. A continuous stirred tank membrane reactor (CST-MR) was developed securing good reactor productivity and high biocatalyst stability. Both production systems are promising, represent two different alternatives and can be further optimized and scaled up for the interests of the industry.

Thesis (PhD)--Stellenbosch University, 2004.
ENGLISH ABSTRACT: Yeasts produce a broad range of aroma-active volatile esters and higher alcohols during alcoholic fermentation. Some of these esters and higher alcohols are important for the fruity flavors and therefore the final quality of wine and other fermented beverages. Esters are produced and hydrolyzed by alcohol acetyltransferases and esterases, respectively. In yeast, ester-synthesizing activities are represented by two alcohol acetyltransferases encoded by the ATFI and ATF2 genes, and by an ethanol hexanoyl transferase encoded by the EHTI gene. Atfl p and Atf2p appear responsible for the production of ethyl acetate and isoamyl acetate, while Ehtl p synthesizes ethyl hexanoate from ethanol and hexanoyl-CoA. Although a fair amount of information is available regarding the ATF 1 gene, limited information is available on the remaining alcohol acetyltransferases. Only two genes that code for esterases have been identified in yeast, namely lAHI and TIPI. It has also been shown that the balance between alcohol acetyltransferases and esterases is important for the net rate of ester accumulation. Higher alcohols are synthesized from the a-keto-acids in the branched-chain amino acid metabolic pathway by decarboxylation and reduction. The transamination of the amino acid to the respective a-keto-acid is catalyzed by mitochondrial and cytosolic branched-chain amino acid transferases, which are encoded by the BATI and BAT2 genes, respectively. In recent years, a strong scientific and industrial interest in the metabolism of flavoractive compounds has emerged, but information regarding the roles of specific enzymes and the physiological relevance of their metabolism remains limited. The aim of this project was to investigate the physiological and metabolic consequences of changes in the expression levels of some of the key enzymes involved in aroma compound production. The consequences of these changes on the chemical composition and the fermentation bouquet of wines and distillates were also investigated. The first part of the section on the results in this dissertation reports on the role and relative importance of the Saccharomyces cerevisiae enzymes involved in ester metabolism, namely Atflp, Atf2p, Ehtlp, Iahlp and Tiplp. The corresponding genes were overexpressed in a laboratory strain of S. cerevisiae, BY4742, and in a widely used commercial wine yeast strain, VIN13. Table wine and base wines for distillation were prepared with these VIN13 transformed strains. The ester concentrations and aroma profiles of the wines and distillates were analyzed and compared. The data indicated that the overexpression of ATF 1 and ATF2 increased the concentrations of ethyl acetate, isoamyl acetate, 2-pheylethyl acetate and ethyl caproate, while the overexpression of JAHI resulted in a significant decrease in the concentrations of ethyl acetate, isoamyl acetate, hexyl acetate and 2-phenylethyl acetate. The overexpression of EHTI resulted in a marked increase in the concentrations of ethyl caproate, ethyl caprylate and ethyl caprate, while the overexpression of TJP1 did not decrease the concentrations of any of the esters. In most cases, there was a correlation between the increase in esters and the decrease in higher alcohols. The data suggest that yeast balances the amount of different esters produced through alcohol acetyltransferases and esterases, and that, in some cases, these enzymes appear to overlap in function and/or influence each other's activity. In the second part of the results section, the consequences of the deletion and the overexpression of two genes, BATl and BAT2, which encode transaminases that contribute to the metabolism of higher alcohols, were investigated. The genes were both disrupted in a S. cerevisiae BY4742, and overexpressed in both this laboratory strain and in the VIN13 wine yeast strain. The effects of these modifications on the general physiology of the corresponding yeast strains and on higher alcohol metabolism were assessed in a range of growth conditions, including aerobic and anaerobic growth conditions, in the presence of glucose or raffinose as sole carbon source and growth in the presence of various concentrations of amino acids. Table wine and base wines for distillation were prepared with the modified industrial strains and the concentrations of the higher alcohols and the aroma profiles of the wine and distillates were analyzed and compared. Batl deletion seemed to be lethal under the conditions that were created, and therefore only the bat2!:!.strain, together with the BATI and BAT2 overexpression strains, were investigated. These modifications did not appear to significantly affect the general physiology of the strains. The results obtained indicated that the overexpression of BATI increased the concentrations of isoamyl alcohol and isoamyl acetate, and, to a lesser extent, the concentrations of isobutanol and isobutyric acid. The overexpression of the BAT2 gene resulted in a substantial increase in the levels of isobutanol, isobutyric acid and propionic acid production, and a modest increase in the level of propanol and isovaleric acid. Interestingly, the overexpression of BAT2 led to a decrease in isoamyl alcohol and isoamyl acetate concentrations. Sensory analyses indicated that the wines and distillates produced with the strains in which the BATl and BAT2 genes were overexpressed had more fruity characteristics (peach and apricot aromas) than the wines produced by the wild-type strains. This study offers new prospects for the development of wine yeast starter strains with optimized ester and higher alcohol-producing capability that could assist winemakers in their efforts to consistently produce wine to definable specifications and styles and a predetermined flavor profile.
AFRIKAANSE OPSOMMING: Gedurende fermentasie produseer giste 'n wye verskeidenheid vlugtige aromatiese esters en hoër alkohole. Sommige van hierdie esters en hoër alkohole is belangrik vir die vrugtige geure en dra dus by tot die finale kwaliteit van wyn en ander gefermenteerde drankies. Esters word onderskeidelik deur alkoholasetieltranferases en esterases geproduseer en gehidroliseer. In giste word die ester-sintetiserende aktiwiteite deur twee alkoholasetieltransferases verteenwoordig wat deur die ATFI-en ATF2-gene, asook 'n etanolheksanoïeltransferase wat deur die EHTl-geen, gekodeer word. Dit blyk dat ATFlp en ATF2p verantwoordelik is vir die produksie van etielasetaat en isoamielasetaat, terwyl Ehtl p-etielheksanoaat vanaf etanol en heksanoïel-CoA sintetiseer. Alhoewel daar 'n redelike hoeveelheid inligting t.o.v die ATF I-geen beskikbaar is, is daar weinig inligting oor die res van die aloholasetieltransferases. Slegs twee gene wat vir esterases kodeer, is in gis geïdentifiseer, naamlik IAHI en TIPI. Daar is ook bewys dat 'n balans tussen die alkoholasetieltransferases en esterases baie belangrik is vir die netto-tempo van ester-akkumulasie. Hoër alkohole word gesintetiseer vanaf a-keto sure in die vertakte-ketting aminosuur metaboliese pad deur dekarboksilasie en reduksie. Die transaminasie van die aminosuur na die onderkeidelike a-ketosuur word deur vertakte-ketting aminosuur transferases, geleë in die mitochondrion en sitosol, en gekodeer deur BATl- en BAT2-gene, gekataliseer. In die laaste paar jare het daar 'n sterk wetenskaplike, asook industrïele, belangstelling in die metabolisme van aroma-aktiewe komponente te voorskyn gekom, maar inligting in verband met die rol van spesifieke ensieme en die fisiologiese belangrikheid van hul metabolisme is egter beperk. Die doel van hierdie projek was om die fisiologiese en metaboliese gevolge van veranderinge in die ekspressievlakke van sommige sleutelensieme betrokke by aromakomponent-produksie te ondersoek. Die gevolge van hierdie veranderinge op chemiese vlakke, asook hoe die fermentasie-aroma van die wyne en distillate beïnvloed word, is ook bestudeer. Die eerste gedeelte van die resultate rapporteer oor die rol en relatiewe belangrikheid van die Saccharomyces cerevisiae-ensieme betrokke by estermetabolisme, naamlik Atfl p, Atf2p, Ehtlp, Iahlp en Tiplp. Die gene was ooruitgedruk in 'n laboratoriurnras van S. cerevisiae, BY4742, asook in 'n kommersïele wyngisras, VIN13. Tafelwyne en basiswyne vir distillasie is gemaak met die getransformeerde VIN13-rasse. Die esterkonsentrasies en aromaprofiele van die wyne en distillate is ontleed en vergelyk. Die data het gewys dat die ooruitdrukking van ATFI- en ATF2-gene 'n verhoging in etielasetaat, isoamielasetaat, 2-fenieletielasetaat en etielkaproaat veroorsaak het, terwyl ooruitdrukking van !AHI 'n betekenisvolle afname in etielasetaat-, isoamielasetaat-, heksielasetaat- en 2-fenieletielasetaat-konsentrasies veroorsaak het. Die ooruitdrukking van EHTI het 'n duidelike verhoging in etielkaproaat, etielkaprilaat en etielkapraat veroorsaak en die ooruitdrukking van TIPIhet geen van die esterkonsentrasies verander nie. In die meeste gevalle was daar nie 'n korrelasie tussen die toename in esters en afname in hoër alkohole nie. Die data stelook voor dat die gis 'n balans tussen die verskillende esters handhaaf deur middel van die alkoholasetieltrasferases en esterases, en in sommige gevalle blyk dit dat die ensieme dieselfde funksies het en/of mekaar se aktiwiteit beïnvloed. In die tweede gedeelte van die resultate is die oorsake van delesie en ooruitdrukking van twee gene, BAT1 en BAT2, wat kodeer vir transaminases wat tot hoër alkohol metabolisme bydra, bestudeer. Die gene is uitgeslaan in S. cerevisiae BY4742 en ooruitgedruk in BY4742 en in die wyngisras VIN13. Die effekte van hierdie modifikasies op die algemene fisiologie van die verskillende gisrasse en op hoëralkoholmetabolisme is onder 'n verskeidenheid kondisies bestudeer, naamlik aërobies en anaërobiese groeikondisies, in die teenwoordigheid van glukose of raffinose as die enigste koolstofbron, asook in die teenwoordigheid van 'n verskeidenheid konsentrasies aminosure. Tafelwyne en basiswyne vir distillasie is gemaak met die gemodifiseerde industrïele rasse en die konsentrasies van die hoër alkohole en aromaprofiele van die wyne en distillate is ontleed en vergelyk. Bat1-delesie was dodelik onder die kondisies, daarom is slegs die batlts-tes tesame met die BAT1 en BAT2 wat in die rasse ooruitgedruk is, bestudeer. Die modifikasies het nie 'n beduidende effek op die algemene fisiologie van die rasse getoon nie. Die data het wel getoon dat die ooruitdrukking van BAT1 'n verhoging in isoamielalkohol- en isoamielasetaatkonsentrasies, en tot 'n mindere mate isobutielalkohol- en isobottersuur-konsentrasies, veroorsaak het. Die ooruitdrukking van BAT2 het 'n beduidende toename in isobutanol-, isobottersuur- en propioonsuurkonsentrasies en 'n kleinere toename in propanol- en isovaleriaansuur veroorsaak. Die ooruitdrukking van BAT2 het ook gelei tot 'n afname in isoamielalkohol- en isoamielasetaatkonsentrasies. Sensoriese analises het getoon dat die wyne en distillate wat geproduseer is met die rasse waarin die BAT1 en BAT2 gene ooruitgedruk is meer vrugtige eienskappe (perske- en appelkoos-aromas) getoon het as die wyne wat deur die wildetipe rasse geproduseer is. Die studie lewer nuwe vooruitsigte vir die ontwikkeling van wyngiste met geoptimiseerde ester en hoër alkohol produserende eienskappe wat die wynmakers in staat kan stelom wyne te produseer met gedefinieerde spesifikasies en style en 'n voorafbepaalde aromaprofiel.

Estery and woody flavour notes are important characteristics of distilled spirit flavour. It has been reported for malt whisky that the estery character of mature whiskies typically declines relative to that of the new make spirit, even though the analytical concentrations of esters remain broadly constant. One potential explanation for this observation would be a sensory interaction between mature and estery characters. The work described in this thesis was designed to test this hypothesis and to further explore the nature of the congeners responsible for eliciting these characteristics across different spirit types, as influenced by their maturation conditions (time, temperature, cask provenance etc.). In the research described in Chapter 2, four pairs of non-mature and mature spirits (tequila, bourbon and 2 malt whiskies) were characterized by instrumental analysis with the aim of defining the key aroma compounds that determined the mature character in each spirit. According to PLS analysis of the full data set, concentrations of 17 congeners were positively correlated with ageing time and might thus influence the mature character of the aged spirits. In Chapter 3, the same eight spirit samples were analysed by GC-Olfactometry using the AEDA (aroma extraction dilution analysis) approach. Aged spirits presented a more complex aroma than new make spirits, and contained more compounds with the highest FD-factors. Whilst a full GC-O characterisation was completed, the main focus was on identifying compounds which contributed to the estery and woody/mature characters of each spirit. In Chapter 4 we attempted to reproduce these characters for each spirit through aroma recombination, based on blends of the odiferous compounds identified at high FD factors and their analytical concentrations in the actual samples as reported in Chapter 3. It soon became apparent that relatively simple mixtures of esters on the one hand and maturation-linked compounds on the other did not adequately reproduce the nature of these characteristics in the spirits themselves. This implied either that our analysis had missed some significant compounds contributing to these characteristics, or that the complexity of the full spirit matrix is required to give the groups of compounds the nuanced flavour that they lacked in isolation. The latter hypothesis was tested by adding in additional blends of compounds to increase the complexity of the recombinant aroma mixtures. It was concluded that the authenticity of the aroma blends overall was improved by both the addition of a cocktail of ‘low boiling compounds’ (those analysed by a separate direct injection GC technique) and the introduction of a ‘structuring’ compound (ethyl hexadecanoate) at a concentration that would cause agglomeration within the whisky (micellar structures) thus influencing aroma partitioning and release. It was concluded that these modifications produced recombinant aromas which were close enough to the authentic spirit characters to use them in sensory interaction studies (Chapter 5). As opposed to interaction effects there was simply a tendency for the woody/mature characters to suppress the corresponding estery character of mature spirits, particularly at the higher concentrations of added wood extractives. Because the woody/mature compounds which characterised maturation were broadly similar across the spirit types, but differed in concentration according to the maturation conditions, we decided finally to investigate the extraction kinetics of wood-derived compounds from oak sticks as a function of ageing time, temperature, spirit type and alcohol content (Chapter 6). Temperature and alcohol content were the most significant factors that determined the extraction rate and final concentrations of all 18 wood-extractive compounds (P < 0.05) analysed. Not surprisingly, extraction rates increased with increasing temperature, but the trend in terms of alcoholic strength depended on the particular compound. Overall this thesis has improved knoEstery and woody flavour notes are important characteristics of distilled spirit flavour. It has been reported for malt whisky that the estery character of mature whiskies typically declines relative to that of the new make spirit, even though the analytical concentrations of esters remain broadly constant. One potential explanation for this observation would be a sensory interaction between mature and estery characters. The work described in this thesis was designed to test this hypothesis and to further explore the nature of the congeners responsible for eliciting these characteristics across different spirit types, as influenced by their maturation conditions (time, temperature, cask provenance etc.). In the research described in Chapter 2, four pairs of non-mature and mature spirits (tequila, bourbon and 2 malt whiskies) were characterized by instrumental analysis with the aim of defining the key aroma compounds that determined the mature character in each spirit. According to PLS analysis of the full data set, concentrations of 17 congeners were positively correlated with ageing time and might thus influence the mature character of the aged spirits. In Chapter 3, the same eight spirit samples were analysed by GC-Olfactometry using the AEDA (aroma extraction dilution analysis) approach. Aged spirits presented a more complex aroma than new make spirits, and contained more compounds with the highest FD-factors. Whilst a full GC-O characterisation was completed, the main focus was on identifying compounds which contributed to the estery and woody/mature characters of each spirit. In Chapter 4 we attempted to reproduce these characters for each spirit through aroma recombination, based on blends of the odiferous compounds identified at high FD factors and their analytical concentrations in the actual samples as reported in Chapter 3. It soon became apparent that relatively simple mixtures of esters on the one hand and maturation-linked compounds on the other did not adequately reproduce the nature of these characteristics in the spirits themselves. This implied either that our analysis had missed some significant compounds contributing to these characteristics, or that the complexity of the full spirit matrix is required to give the groups of compounds the nuanced flavour that they lacked in isolation. The latter hypothesis was tested by adding in additional blends of compounds to increase the complexity of the recombinant aroma mixtures. It was concluded that the authenticity of the aroma blends overall was improved by both the addition of a cocktail of ‘low boiling compounds’ (those analysed by a separate direct injection GC technique) and the introduction of a ‘structuring’ compound (ethyl hexadecanoate) at a concentration that would cause agglomeration within the whisky (micellar structures) thus influencing aroma partitioning and release. It was concluded that these modifications produced recombinant aromas which were close enough to the authentic spirit characters to use them in sensory interaction studies (Chapter 5). As opposed to interaction effects there was simply a tendency for the woody/mature characters to suppress the corresponding estery character of mature spirits, particularly at the higher concentrations of added wood extractives. Because the woody/mature compounds which characterised maturation were broadly similar across the spirit types, but differed in concentration according to the maturation conditions, we decided finally to investigate the extraction kinetics of wood-derived compounds from oak sticks as a function of ageing time, temperature, spirit type and alcohol content (Chapter 6). Temperature and alcohol content were the most significant factors that determined the extraction rate and final concentrations of all 18 wood-extractive compounds (P < 0.05) analysed. Not surprisingly, extraction rates increased with increasing temperature, but the trend in terms of alcoholic strength depended on the particular compound. Overall this thesis has improved knowledge of the chemical and sensory changes that accompany spirit maturation and has highlighted some of the factors that cause differences in mature character across spirit types.
Moreover, it concludes that the sensory perception of woody/mature generally suppresses the fresh estery characteristic of new make spirits, even though analytically the esters are still there in similar concentrations. wledge of the chemical and sensory changes that accompany spirit maturation and has highlighted some of the factors that cause differences in mature character across spirit types. Moreover, it concludes that the sensory perception of woody/mature generally suppresses the fresh estery characteristic of new make spirits, even though analytically the esters are still there in similar concentrations.

L'elaboration de boissons alcoolisees par fermentation de produits vegetaux est une pratique tres ancienne qui a connu de reels progres depuis les travaux de pasteur au 19#e siecle. La selection des varietes de raisin, le choix du meilleur moment pour la vigne ainsi que le controle du procede fermentaire, sont autant de facteurs d'amelioration du vin. Pour notre part, nous nous sommes interesses a la phase de fermentation alcoolique qui contribue tant sur le plan quantitatif que qualitatif a la formation des aromes du vin. Bien que ces composes soient tres nombreux, les principaux ont ete identifies et il est admis que sept d'entre eux se partagent plus de la moitie des aromes dans le vin : cinq alcools superieurs (propanol-1, isobutanol, 2-methyl et 3-methyl butanol-1, phenyl-2 ethanol) et deux esters (acetate d'ethyle et lactate d'ethyle). Les parametres, influant sur la synthese de ces composes, ont fait l'objet de nombreuses etudes mais a des resultats contreverses. D'autre part toutes les investigations entreprises, ont porte sur des evaluations de bilans entree-sortie et aucune ne decrit de facon precise les cinetiques de production. Afin de combler partiellement cette lacune et grace a un chromarographe de type espace-tete, nous avons etabli les cinetiques de production des aromes, ce qui constitue l'originalite de notre travail. Nous avons egalement souhaite preciser par notre etude, le role de parametres importants sur les cinetiques de production. En effet, nous nous sommes interesses a l'effet de la : temperature, la souche de levure, le taux de biomasse initial et a la composition du milieu en acides amines. Par un souci de reproductibilite, nous avons, dans une premiere etape, realise nos experiences sur un milieu modele, puis dans un deuxieme temps, nous avons valide les resultats obtenus sur un mout de raisin naturel : du mauzac.

La réalité sensorielle de l'existence d'un arôme fruité spécifique aux vins rouges a été démontrée par le biais de dégustations affranchies de toute subjectivité liée aux caractéristiques visuelles des vins. Ces dégustations ont également mis en évidence une typicité olfactive des vins rouges de Bordeaux. L'étude de la genèse de cet arôme fruité propre aux vins de Vitis vinifera L. Cv. Merlot et Cabernet-Sauvignon a ensuite révélé le rôle clé de la fermentation alcoolique d'une part et de la présence de constituants pelliculaires extractibles à l'éthanol d'autre part. L'intensité de l'expression fruitée est par ailleurs modulée par la présence de composés soufrés - thiols en particulier - et/ou de leurs précurseurs. La mise au point d'une méthode de fractionnement par HPLC d'extraits aromatiques a permis d'isoler le caractère fruité des vins rouges dans des fractions spécifiques, préservant fidèlement les nuances aromatiques correspondantes. Suite à l'étude GC-olfactométrique des fractions ainsi obtenues, de nombreuses zones odorantes, spécifiques ou non des vins rouges, ont pu être mises en évidence. Les composés aromatiques responsables de vingt d'entre elles ont été identifiés. Ils correspondent à des esters éthyliques et acétates, dont les teneurs dans des vins rouges ainsi que les seuils de perception dans la même matrice ont été déterminés. Ces composés ne présentent pas d'impact direct sur l'arôme fruité des vins rouges. Des tests de reconstitutions aromatiques ont en revanche clairement établi leur impact en mélange, reflet d'interactions perceptives entre ces composés et pouvant conduire à des nuances fruitées de type fruits rouges ou fruits noirs.

Thesis (MSc)--University of Stellenbosch, 2002.
ENGLISH ABSTRACT: No single chemical constituent can be accredited with giving wine and brandy their overall aroma and flavour. The aroma and flavour of wine and brandy are rather attributed to a number of chemical constituents reacting together and it is these reactions that give the beverage its character. Certain chemicals within wine and brandy do, however, make larger contributions to the flavour. These include the esters, terpenes and volatile acids, although others also exist. Esters are a large group of volatile compounds with variable aroma and flavour characteristics, including banana-like (isoamyl acetate), apple-like (ethyl caproate) and chemical/solvent-like (ethyl acetate). Esters are produced as secondary metabolites during the conversion of sugar to ethanol and are formed when an alcohol binds with a fatty acid. Chemically, ester metabolism is well documented and understood; however, much work still needs to be done on a genetic level. The yeast strain used during fermentation is one of the most important factors contributing to the type and quantity of esters produced. This is due to differences in genetic makeup. The metabolism of esters is controlled largely on a genetic level, with numerous genes being involved. The alcohol acetyltransferase genes are involved in ester anabolism, whilst esterase genes are involved in ester catabolism. Esterases have a negative effect on the overall level of esters within an alcoholic beverage, as they are capable of reducing the number of esters and are thus capable of altering the beverage's aroma and flavour profile. The IAH1 and the TIP1 gene products are believed to encode for two such esterases. The objective of this study was to investigate the contribution of the IAH1 and TIP1 genes to the level of esters in both wine and brandy. This was accomplished by using two approaches. Firstly, the above genes were disrupted using a polymerise chain reaction (PCR)-generated disruption cassette homologous to either the IAH1 or the TIP1 gene. These cassettes were integrated into the industrial wine yeast, Saccharomyces cerevisiae strain VIN13. The integrations were verified by Southern blot analysis to produce yeasts VIN13-~IAH1 and VIN13-~TIP1; however, only a single copy of each was disrupted. Secondly, the IAH1 and the TIP1 genes were cloned from S. cerevisiae using PCR into plasmid pj between the phosphoglycerate kinase gene (PGK1) promoter and terminator, producing plasmids pJ-IOE1 and pJ-TOE1. The PGK1 promoter has previously been shown to constitutively express genes at high levels. These new constructs were then used as template for PCR to produce two overexpression cassettes, one for IAH1 and the other for TlP1. These cassettes were integrated into S. cerevisiae VIN13 and verified by Southern blot analysis to produce strains VIN13-IOE1 and VIN13-TOE1. The above yeast strains including VIN13 were used for the production of wines and base wines from Colombard must. Reverse-transcriptase (RT-PCR) confirmed that the VIN13-IOE1 and VIN13-TOE1 strains overexpressed the appropriate gene at a higher level than the control VIN13 strain. The VIN13-AIAH1 disrupted strain showed no difference in expression level to that of the control strain, whilst VIN13-ATIP1 showed lower levels of expression than that of the control strain. VIN13-IOE1 behaved as expected, with a decrease of between 30% and 60% in the total ester level in the wine and base wine respectively, a 30% decrease in the total acid level and no change in the higher alcohol level. The VIN13-AIAH1 strain showed no difference to the control wine, most likely as this strain still expressed the IAH1 gene at levels consistent with the control strain. VIN13-TOE1 behaved in an unexpected manner - instead of hydrolysing esters, it appeared to produce them. This increase in the total ester level was most noticeable during distillation, when a 20% increase took place. Another unexpected occurrence was a large decline in the total acid level, with acetic acid being the most significant contributor, decreasing by up to 78%. This is a very favourable finding, as acetic acid is a known spoilage molecule and is a cause of sluggish/stuck fermentations. VIN13-ATIP1 behaved in an opposite manner to VIN13-TOE1, with higher total acid levels and slightly decreased total ester levels, especially during distillation. Neither affected the total higher alcohol levels. Sensorially, the only significant difference in the wine samples was for the fruity flavour. A panel of judges distinguished that VIN13-TOE1 was fruitier than the other wines, with VIN13-ATIP1 being the least fruity. This study again proves the significant impact that a single gene can have on the chemical makeup of wine and brandy. The relatively simple genetic alteration of an organism can drastically change and improve not only the organoleptic properties of the organism, but its viability as well. These alterations can produce more favourable organisms with more desirable characteristics for the fermenting beverage industry to produce products of higher quality and better suitability.
AFRIKAANSE OPSOMMING: Geen chemiese komponent kan uitgesonderword as die produseerder van aroma en geur in wyn of brandewyn nie. Die aroma en geur van wyn en brandewyn word eerder toegeskryf aan die interaksie tussen 'n groot aantal chemiese komponente om aan die drank sy karakter te gee. Enkele van hierdie chemiese komponente sluit in esters, terpene en vlugtige sure, om maar 'n paar te noem. Esters is "n groot groep van vlugtige verbindings wat beskik oor 'n verskeidenheid van aroma- en geurkenmerke, soos piesangagtig (isoamielasetaat), appelagtig (etielkaproaat) en chemies/oplosmiddelagtig (etielasetaat). Esters word as sekondêre metaboliete geproduseer wanneer suikers na etanolomgeskakel word en word gevorm wanneer "n alkohol met "n vetsuur verbind. Estermetabolisme is chemies goed beskryf en verstaan, maar op "n genetiese vlak is daar nog heelwat aspekte wat nagevors moet word. Die gisras betrokke gedurende fermentasie word beskou as een van die grootse bydraes tot die tipe en die hoeveelheid esters wat geproduseer word. Dit word toegeskryf aan verskille in die genetiese saamestelling van die gisras. Ester metabolisme word grootliks deur genetiese faktore beheer en verskeie gene is betrokke. Dit is hoofsaaklik die alkoholasetieltransferasegene wat vir esterkatabolisme verantwoordelik is, terwyl die esterasegene vir esteranabolisme verantwoordelik is. Esterases het 'n negatiewe effek op die totale estervlak binne alkoholiese dranke deurdat hulle in staat is om die aantal esters drasties te verminder en sodoende die drank se aroma- en geurprofiel te verander. Daar is voorgestel dat die IAH1- en die TlP1-geen produkte is wat vir twee sulke esterases kodeer. Die doel van hierdie studie was om die IAH1- en die TIP1-gene se bydrae tot die totale estervlak in wyn en brandewyn te ondersoek. Dit is deur twee benaderings uitgevoer. Eerstens is die bogenoemde gene d.m.V. disrupsiekassette wat homoloog aan die IAH1- of die TlP1-gene was, uitgeslaan. Die disrupsiekassette is deur die polimerasekettingreaksie (PKR) geproduseer. Hierdie kassette is in die industriële wyngis, Saccharomyces cerevisiae VIN13, geïntegreer. Die integrasies is deur Southernkladanalise bevestig en het die giste VIN13-~IAH1 en VIN13-~TIP1 gelewer. Net 'n enkele kopie van elke geen is egter uitgeslaan. Tweedens is die IAH1- en TIP1-gene d.m.V. PKR vanaf S. cerevisiae binne in plasmied pJ gekloneer, tussen die fosfogliseraatkinasegeen (PGK1) se promotor en termineerder, om plasmiede pJ-IOE1 en pJ-TOE1 te produseer. Die PGK1-promotor is al tevore geïdentifiseer as "n hoë-vlak konstitutiewe uitdrukker van gene. Hierdie twee nuwe konstrukte het vervolgens gedien as templaat vir PKR om twee ooruitdrukkingskassette, een vir IAH1 en die ander vir TIP1, te produseer. Hierdie kassette is in S. cerevisiae VIN13 geïntegreer en bevestig deur Southernkladanalise. Hierdie integrasies het die giste VIN13-IOE1 en VIN13-TOE1 geproduseer. All die nuwe gisrasse, tesame met VIN13, is gebruik vir die produksie van wyne sowel as rebatwyne vanaf Colombard-mos. Omgekeerde-transkriptase polimerasekettingreaksie (OT-PKR) het bewys dat die VIN13-IOE1 en VIN13-TOE1 rasse die geskikte geen ooruitgedruk het, met hoêr vlakke as van die kontrole VIN13-ras. Dit het ook aangedui dat die VIN13-i\IAH1-ras, waarvan die geen uitgeslaan was, geen verskil in uitdrukking gehad het in vergelyking met die kontroleras nie, terwyl VIN13-i\TIP1 'n lae uitdrukkingsvlak getoon het. VIN13-IOE1 het teen verwagting opgetree, met 'n afname van tussen 30% en 60% in die totale estervlak in beide die wyne en rebatwyne. 'n Afname van 30% in die totale suurvlak, asook geen waarneembare verskil in die hoêr alkoholvlak, in vergelyking met die kontroleras, is ook opgemerk. Die VIN13-i\IAH1-ras het glad nie van die kontroleras verskil nie, heel waarskynlik omdat hierdie ras die IAH1-geen teen dieselfde vlak as die kontroleras kon uitdruk. Die VIN13-TOE1-ras het teen verwagting opgetree deurdat dit esters geproduseer het i.p.v. om esters te hidroliseer. Hierdie toename in die totale estervlak is die meeste waarneembaar tydens distillasie, met tot 'n 20% toename. Nog 'n onverwagte effek was die groot afname in die totale suurvlak. met asynsuur wat die betekenisvolste bydrae gelewer het deurdat dit 'n afname van tot 78% getoon het. Hierdie bevinding is baie voordelig, aangesien asynsuur, 'n bekende bederfmolekuul, veral vir slepende/gestaakte fermentasies verantwoordelik is. VIN13-i\TIP1 het op die teenoorgestelde wyse opgetree as VIN13-TOE1, met 'n hoêr totale suurvlak en 'n klein afname in die totale estervlak. Weereens is dit meer gedurende distillasie waargeneem. Beide rasse het egter geen effek op die hoêr alkoholvlak gehad nie. Die proepaneel het, met betrekking tot die vrugtige geur, een betekenisvolle geurverskil tussen die wyne gevind. VIN13-TOE1 was meer vrugtig as al die ander wyne en VIN13-i\TIP1 was die minste vrugtig. Die studie het weereens bewys dat 'n enkele geen 'n betekenisvolle effek op die chemiese samestelling van wyn en brandewyn kan hê. Die relatief eenvoudige genetiese verandering van 'n organisme kan die organoleptiese eienskappe asook die lewensvatbaarheid van "n organisme, drasties verander en verbeter.

Les fermentations alcoolique (FA) et malolactique (FML) sont deux étapes importantes de la vinification en rouge dans l’établissement de l’arôme fruité des vins. Afin d’étudier l’importance relative des microorganismes fermentaires, nous étudions l’influence de six couples levures/bactéries lactiques (BL) - trois souches de levures, deux de BL - sur la modulation des notes fruitées de différents vins rouges de Bordeaux. Une première approche analytique montre l’influence prédominante de la souche de levures sur la concentration de plus de 70 marqueurs potentiels de la note fruitée. L’étude particulière des esters montre que l’effet levures observé dès la fin de la FA persiste au cours du temps malgré la FML et les modifications engendrées par le vieillissement du vin. L’étude sensorielle conforte l’influence majeure des levures sur la modulation de l’arôme fruité des vins rouges à différents temps d’élevage. Néanmoins, les résultats obtenus suggèrent l’implication d’autres composés aromatiques dans la modulation de la note fruitée des vins, non quantifiés dans la première partie de cette étude. Un travail de fractionnement d’extraits de vin par HPLC permet par la suite l’identification d’une fraction d’intérêt impliquée dans des variations aromatiques liées à la souche de levures. L’analyse de cette fraction par chromatographie en phase gazeuse n’a pas permis d’identifier le ou les composés impliqués. Nous avons néanmoins mis en évidence une thiophénone qui pourrait agir en tant que masque de l’arôme fruité, ainsi qu’un ester hydroxylé qui pourrait s’avérer être un marqueur intéressant de l’activité bactérienne et dont l’effet exhausteur de notes fruitées est également envisagé comme perspectives
Alcoholic (AF) and malolactic (MLF) fermentations are important steps in red winemaking for the revelation of wine fruity aroma. To investigate the relative importance of fermentative microorganisms, we studied the influence of six yeasts/lactic acid bacteria (LAB) combination - three yeast strains, two LAB - on Bordeaux red wines fruity notes modulation. A first analytical approach showed the predominant influence of yeast strain on the concentration of more than 70 potential fruity note markers. Special study of esters showed a yeast strain effect since the end of FA that persists over time, despite MLF and changes caused by wine aging. Sensory studies also highlighted the major influence of yeasts on red wines fruity aroma modulations at different aging steps. Nevertheless, results suggested the role of other aromatic compounds in fruity note modulation, not quantified in the first part of this study. The study of fractions made by HPLC with wine organic extracts enables the identification of an interested fraction involved in aromatic variations related to the yeast strain. Analysis of this fraction by gas chromatography has not allowed identifying compounds involved in these organoleptic variations. However, we highlighted a thiophenone that could act as a mask of fruity aroma and a hydroxylated ester that could be an interesting marker of bacterial activity. Its role as enhancer of fruity esters aroma is also considered

L’objectif de cette thèse est d’étudier le rôle de la fermentation malolactique (FML) sur l’arôme fruité des vins rouges. Les bactéries lactiques (BL) modifient la composition du vin mais il n’existe pas de consensus concernant spécifiquement cette famille aromatique. Contrairement aux idées empiriques sur la FML, ce travail a démontré l’absence à court terme d’un " masque lactique ", cependant l’apparition d’une telle interaction olfactive pourrait être plus tardive. Par contre, il est montré l’existence d’un masque proche de la note de réduction, de type fumé/grillé, dont la caractérisation n’a pas été effectuée dans cette étude.Le suivi des principaux marqueurs fruités du vin (70 molécules) a été rendu possible par le développement des méthodes d’analyse chromatographique en phase gazeuse couplée à la microextraction sur phase solide (esters, C13-norisoprénoïdes, lactones, thiols). En particulier, une " base de données esters " (32 composés) a rendu plus robuste l’ensemble des variations constatées au cours du développement des BL. En effet, les modifications des teneurs en esters sont démontrées comme un processus majeur de la balance de la note fruitée au cours de la FML. Cette fermentation permet à court terme, aussi bien la synthèse que l’hydrolyse des esters grâce aux activités estérases et, à plus long terme, la formation tardive d'esters éthyliques d'acides branchés issus du catabolisme de certains acides aminés. La spécificité des estérases vis-à-vis de la nature et de la longueur de la chaîne carbonée des esters est mise en évidence, ainsi que l'importance de la disponibilité des substrats, liée en partie à l'activité des levures.L’étude de l’influence des souches de BL et de la co-inoculation levures/bactéries a permis de confirmer le rôle clé des interactions entre les microorganismes, ainsi que l’importance de la composition de la matrice vin
The aim of this thesis is to study the role of malolactic fermentation (MLF) on the fruity aroma of red wines. Lactic acid bacteria (LAB) modify wine composition but there is no consensus concerning this aromatic group specifically. In opposition to empirical ideas on MLF, this work has demonstrated the absence, in short-term, of a “lactic-mask” although this kind of olfactory interaction may still occur in a later stage of wine development. Nevertheless, the existence of a smoked/toasted reduction-like mask note was proved. Its characterization has not been done in this work. The survey of the main fruity markers of wine (70 compounds) was made possible by the development of several gas chromatography coupled with solid-phase microextraction analytical methods (esters, C13-norisoprenoids, lactones, thiols). In particular, the creation of an “ester database” (32 compounds) has improved the detection of variations during LAB development in terms of analysis robustness. In fact, changes on esters contents are proved to be responsible for a major part of fruity notes evolution during MLF. Initially, this fermentation allows both synthesis and hydrolysis of esters as a consequence of esterase activity. Moreover, it promotes late-production of ethylic esters through the catabolism of certain aminoacids. Esterases specificity towards nature and size of the esters carbon chain is pointed out along with substrates availability, partially related to yeast activity.The study of the influence of both LAB strains and yeast/bacteria co-inoculation has confirmed microorganisms interactions and wine matrix composition to be of the great importance

L’expression aromatique fruitée des vins rouges a été le sujet de nombreuses études qui démontrent qu’au moins une composante de cette expression est le reflet d’interactions perceptives impliquant des esters. La plupart des travaux concernant les interactions perceptives jusqu’à ce jour se sont avérés descriptifs, très peu ayant cherché à déterminer leurs origines. Dans ce but, un outil analytique a été développé afin d’apprécier les changements de volatilité d’esters représentatifs de l’arôme fruité des vins rouges. Ainsi, les coefficients de partage de 9 esters ont pu être déterminés aussi bien dans une solution hydroalcoolique que dans un vin rouge désaromatisé. L’application de cet outil analytique aux interactions perceptives préalablement mises en évidence a permis d'observer des changements de volatilité des esters lors de leur mise en mélange avec d'autres composés volatils en solution. Ces changements de volatilité, synonymes de potentiels effets pré-sensoriels, vont dans le même sens que ceux observés lors de l’analyse sensorielle. L’utilisation d’un verre de dégustation possédant deux compartiments a permis de mettre en lumière le fait que certaines modifications sensorielles pouvaient être expliquées, pour partie au moins, par des effets pré-sensoriels. L'impact olfactif de 5 alcools supérieurs ainsi que de 15 composés issus du bois de chêne a pu être démontré grâce à de nombreuses reconstitutions aromatiques, et leur rôle de masquage de l’arôme fruité des vins rouges a pu être souligné. Le calcul des coefficients de partage des esters a permis de montrer que des changements de volatilité ont lieu au sein de la solution. Ces modifications peuvent être corrélées aux résultats obtenus lors de l’analyse sensorielle. Ainsi, il est possible d’expliquer, en partie, les effets de masquage de l’arôme fruité observés grâce aux seuils de détection et aux profils sensoriels, du fait de la diminution de la présence d’esters dans l’espace de tête venant stimuler le dégustateur. Globalement, nos travaux ont permis de mettre en évidence que la mise en mélange en solution de composés volatils pouvait se traduire par la modification de la volatilité des constituants du mélange et que certaines de ces interactions pré-sensorielles pouvaient conditionner l'expression aromatique fruitée due aux esters
A lot of studies highlighted the perceptual role of esters in fruity aromatic expression of red wines, demonstrating that at least partially it was due to perceptive interactions. Indeed, a lot of synergistic and masking effects have been brought to light in the past. However, the origin of these interactions remains unknown, although some authors suggested several levels where they can take place. In this goal, an analytical tool was developed to study the possible occurrence of esters volatility modifications. The application of this tool allowed determining partition coefficients of 9 esters in dilute alcohol solution and in dearomatized red wine. Thanks to perceptive interactions previously demonstrated by various authors, the application of this analytical tool highlighted modifications of esters volatility when compounds were mixed together in the solution. These modifications support the observations made with sensory analysis, indicating the existence of pre-sensorial effects. The use of a new tool consisting in a tasting glass with 2 compartments, reveals that these volatility changes may led to true sensorial modifications. Masking effect of fruity aroma due to 5 higher alcohols but also 15 wood by-products was highlighted using various aromatic reconstitutions. Esters partition coefficients calculation showed volatility modifications from the matrix to the gas phase. These data may be correlated with sensorial analysis results. Thus, it is possible to explain, at least partially, fruity aroma masking effect highlighted through detection threshold and sensory profile thanks to decrease in esters presence in headspace, and so a decline of taster’s olfactory stimulation. To conclude, our work showed that the mixture of volatile compounds in solution may result in modification of molecules volatility, and furthermore highlighted that these pre-sensorial interactions may impact fruity aromatic expression related to esters

To develop one porous additive material that can be assembled on cigarette filter, and achieve both functions of reducing hazard substances like tar and providing special aroma and moisture, this study innovatively selects the raw materials like baking-carbonized functional flavors with special aroma and konjac glucomannan (KGM) with properties of water absorption, gelling and film-forming, following the steps of casting into membrane shape, thermostatic crosslinking for strength enhancement and freeze-drying for pore creating to prepare 5 porous composite membranes based on baked-carbonized functional flavors and konjac glucomannan (KGM) were prepared. The composite structure, morphologies on the surface and the cross section, porosity structures including mesoporous and macropore of the porous composite membranes were characterized, the tensile performance and aroma constituents through solid phase microextraction and hazardous substances adsorption as packed into the commercial cigarette were also studied. Results showed that all baked-carbonized functional flavors were carbonized into amorphous, and contained the groups of aromatic components like ≡C-H, C=O, C-O-C (aldehydes, esters), cross-linking achieved the entanglement of KGM segments by deacetylation. When compounded, KGM acted as the framework to build the 3D porous network structure, and the functional flavor powders were wrapped into the KGM layer. All porous composite membranes owned some mechanical strength and the internal porosity over 90%, the mesopores and the macropores were ranged with 5–50 nm and 20–255 µm, respectively, which was satisfied for gas flowing and harmful substances adsorption. Aroma and resulting abundance of extracted constituents changed with timing, which equipped the membranes with a comprehensive aroma feeling. Among them, the membrane prepared with the raw material of dark plum showed a better comprehensive performance, especially it could significantly reduce the contents of harmful HCN and phenol in cigarette during smoking.

Strawberries of such cultivars as “Ducat”, “Honey” and “Polka” were studied to define the content of aromatic volatiles using the methods of highly efficient liquid chromatography. About 49 components were identified in the composition of volatiles of ripe strawberries, namely esters, aldehydes, ketones, furanone, organic acids, aroma compounds, lactones, terpenic compounds, and alkanes. Their shares are the following: esters—12.8–41.8%, aldehydes—5.9–15.9%, ketones—8.7–35.6%, furanone—22.7–24.4%, and organic acids—2.47–21.85%. Depending on a cultivar, typical volatile components of strawberries are ethyl butanoate (10.1–30.65%), trans-2-hexenal (5.31–15.55%), acetoin (8.20–35.67), 2.5–dimethyl-4-methoxy-3(2H)-furanone (mesifurane) (19.08–19.92%) and 2.5-dimethyl-4-hydroxy-3(2H)-furanone (3.43–4.40%). A peculiar feature of volatile compounds of Polka strawberries is the highest ester share—41.77% of total content of volatiles, for Ducat strawberries it would be the share of ketones (35.88%), and for Honey strawberries—γ-decalaktone (12.41%). A high aroma activity of ripe strawberries is recorded on 2.5-dimethyl-4-methoxy-3(2H)-furanone (mesifurane) and 2.5-dimethyl-4-hydroxy-3(2H)-furanone (furaneol). Sweet, caramel flavors are typical for strawberries of the studied cultivars. Strawberries of Polka cultivar have a pronounced aroma.

Whisky is an oak-matured spirit that is produced mainly from malted barley, wheat and maize, although other substrates, including molasses and sorghum are used in certain parts of the world. In cereal substrates the carbohydrates necessary for the production of ethanol are bound up as unfermentable starch which needs to be broken down before fermentation can proceed. To do this, the starch is gelatinized by heating in the presence of water above the critical point of its gelatinization temperature. This gives the saccharifying enzymes access to the polysaccharide polymers, which progressively release fermentable sugars. Parallel proteolytic activities also release amino acids which facilitate yeast growth and are important in the development of flavour-active higher alcohols and esters. After fermentation, which can take as little as 45–55 hours to complete, the contents of the fermenter are transferred to stills for ethanol and volatile flavour recovery. The distillery may operate either in batch or continuous mode and the resulting spirit, which in many producing regions including Ireland and Scotland cannot exceed 94.8% alcohol by volume (ABV), is then diluted with water to maturation strength (typically 63.5% ABV) before being filled into casks for maturation. Whilst overall whisky production takes the order of years, the processes from crop-to-cask only take a few days to execute, with subsequent maturation often being the most protracted part of the production process.

Os co-produtos (cascas e albedo) de maracujá é um resíduo que pode ser aproveitado na forma de farinha. O arroz apresenta característica de hipoalergenicidade, com flavor suave e isento de glúten. Este trabalho teve como objetivo desenvolver formulações de biscoitos tipo cookie sem glúten contendo uma farinha mista extrudada de cascas e albedo de maracujá(Passiflora edulis flavicarpa Degener) e arroz (Oryza sativa L.) e avaliação física. Para os cookies avaliou-se a aceitabilidade sensorial e avaliação física. A análise sensorial do cookies foi realizada com 100 julgadores não treinados, utilizando uma escala hedônica de 9 pontos, mediante a assinatura do Termo de Consentimento Livre e Esclarecido. Para determinar a avaliação física do cookies foi realizado valores médios da massa, volume aparente, volume específico, diâmetro, espessura e fator de expansão. Os resultados obtidos mostram que das três formulações (Fb8, Fb12 e Fb13) desenvolvidas, duas (Fb8 e Fb13) obtiveram aceitação por parte dos julgadores. Os resultados da avaliação física indicaram que o diâmetro e o fator de expansão obtiveram os maiores valores. Portanto, concluí-se que o uso de co-produtos (cascas e albedo de maracujá) na fabricação de farinha para cookies pode ser uma alternativa viável para o consumo por portadores de doença celíaca.

A leishmaniose é uma doença produzida por parasitos protozoários do gênero Leishmania sp e é transmitida ao homem e a outras espécies de mamíferos por insetos vetores denominados flebotomíneos. Apresenta diferentes manifestações clínicas, sendo a Leishmaniose Visceral (LV) a forma mortal se não for tratada. Até hoje, nenhum tratamento foi desenvolvido para esta doença, as alternativas terapêuticas utilizadas são altamente tóxicas e diversos parasitos vem apresentando resistência ao longo dos anos. Por isso, é necessário que se desenvolvam fármacos contra alvos específicos das vias biosintéticas da Leishmania. Este capítulo relata um estudo realizado com 214 extratos de plantas da Mata Atlântica Fluminense para a busca de inibidores da enzima nucleosídeo hidrolase da Leishmania donovani (LdNH). Do extrato das folhas da Urvillea rufescens Cambess (Sapindaceae) foi isolada a tricetina-4´-metoxi-flavona e do extrato das flores de Leandra amplexicaulis DC (Melastomataceae) foram isolados o kaempferol 3-O-α-L-rhamnopyranosido e o kaempferol 3-O-β-D-xylopyranosyl-(1→2)- α-L-rhamnopyranosido. Todos os compostos foram caracterizados estruturalmente por ressonância magnética nuclear (RMN) e espectrometria de massas (EM). No ensaio de inibição da LdNH a tricetina-4´-metoxiflavona apresentou uma concentração inibitória média (CI50) de 1,19 ± 0,26 µM, sendo considerada um bom inibidor desta enzima. Pelo que sabemos, esta é a primeira vez que um flavonoide é descrito como inibidor de NH.

Introdução: O tomilho (Thymus vulgaris L.) é uma planta da família Lamiaceae que compreende cerca de 350 espécies. Muitas das espécies introduzidas no Brasil são plantas medicinais, aromáticas e produtoras de óleos essenciais. Objetivo: Este trabalho tem como objetivo revisar a eficácia do óleo essencial de tomilho na atividade antimicrobiana. Material e métodos: Foram selecionados do site Scopus 7 artigos científicos sobre o tema em estudo com as palavras chaves: Thymus vulgaris, essential oil e anti microbial activity. Resultados: A utilização de óleo essencial de tomilho como agente antifúngico mostrou-se eficiente como inibidores de Aspergillus niger e A. Flavus, dois fungos importantes na indústria de alimentos. Quando estudado o A. niger, o óleo essencial de tomilho reduziu o crescimento fúngico, quando utilizadou-se 2 mL / 18 mL de meio de cultura, embora o total de inibição foi apenas quando 8 mL foi usado. Para inibir crescimento A. flavus, óleo essencial de tomilho reduziu o crescimento micelial a 2,4 e 6 mL. A inibição foi total em 8 mL, isso significa que A. Flavus é sensível ao óleo essencial de tomilho. Este óleo essencial pode ser utilizado como agente antifúngicos, sendo a principal razão para a sua adequação, a sua origem natural, que os consumidores encontrar reconfortante e que é benéfico para o meio ambiente, e possui um risco muito baixo de patógenos que irão desenvolver resistência à mistura de componentes que fazem o óleo com sua aparente diversidade de mecanismos de antifúngicos. O óleo essencial de tomilho mostrou 60% de inibição a concentração 10 mL−1 contra Alternaria brassicicola e apresentou quase 100% de inibição em 1 mL−1 contra a algumas espécies de fungos. O mais prevalente composto de tomilho é o timol (44,60%), sendo o timol e carvacrol são isómeros posicionais com os seus grupos hidroxilo fenólicos em diferentes locais. Conclusão: Estes resultados confirmam o potencial uso do óleo essencial de tomilho na indústria para a preservação de alimentos contra fungos e bactérias, além de aumentar a vida de prateleira de gêneros alimentícios.

Introdução: O Aspergillus flavus é um fungo endêmico do solo, foi identificado como grande produtor de aflatoxina, a contaminação ocorre tanto durante o armazenamento, quanto ainda na fase de plantio. O milho é uma das culturas mais afetadas pela espécie, levando à sua contaminação por aflatoxina, e posteriormente esta é repassada aos animais que o consomem. O uso de fungicidas químicos é amplamente aplicado, no entanto esta prática pode levar à contaminação do grão. O biocontrole vêm como uma alternativa sustentável para substituir os defensivos agrícolas químicos. Objetivos: Na bactéria Bacillus amyloliquefaciens–VR002 (BA) foram identificadas características relacionadas ao seu papel como antagonista de microrganismos patogênicos. Analisou-se a atuação do sobrenadante da bactéria BA-VR002 no crescimento micelial do fungo Aspergillus flavus. Material e métodos: Os experimentos ocorreram no Laboratório de Biotecnologia/UFF-VR. A bactéria foi crescida utilizando 150mL do meio Luria Bertani esterilizado e inoculado com a cultura de BA-VR002 deixando-a crescer por 48h em estufa à 37ºC e 170rpm. Após o tempo de fermentação, o caldo foi centrifugado à 3500rpm, à 4ºC por 30min, filtrado em filtro de 22μm para um erlenmeyer estéril. Este filtrado foi então misturado ao meio ágar LB à 50±10ºC nas proporções de: 0%(v/v); 10%(v/v); 20%(v/v) e 30%(v/v). Este foi vertido em placas de Petri estéreis, após o endurecimento do ágar foram dispostos discos de micélio contendo o fungo A. flavus no centro das placas incubando-as em estufa à 30ºC por 5 dias. Os testes foram feitos em sextuplicata, e a área de crescimento foi calculada por meio do programa ImageJ. Resultados: Os resultados mostram que o percentual de inibição fúngico máximo ocorreu na concentração de 30% de sobrenadante, atingindo 96,4% de inibição do crescimento micelial quando comparado ao controle. Na concentração de 10% a inibição foi de 14,8%, na de 20% foi de 84,6%, confirmando a presença das substancias antifúngicas produzidas pela BA-VR002. Conclusão: O sobrenadante da bactéria B. amyloliquefaciens-VR002 contém substâncias antifúngicas, sendo a concentração de 30% a mais eficaz, mostrando-se como um potencial agente de controle do fungo A. flavus.

Short path distillation and thin film evaporation are generally associated with processing of heat-sensitive oils that require short residence times, high heat and mass transfer coefficients and shorter contact times. Thin film evaporation and short path distillation processes that normally operate at lower temperatures and higher vacuums are commonly used in the processing heat-sensitive oils like omega 3 oils and concentration of omega 3 ethyl esters. These technologies are now also being considered for the removal of processing and environmental contaminants from oils like palm, soybean and other marine oils.Artisan's thin evaporation and the new short path stripping (SPS) technology, with internal multistage-condensing capabilities, take advantage of both principles, namely short path distillation and thin film evaporation. Artisan-SPS system, with no internal moving parts and higher processing capacity, offers several advantages over commercial molecular distillation and deodorizer systems including applications in the removal of contaminants, physical refining and recovery and concentration of omega 3 ethyl esters.This presentation will include results on applications of the new thin evaporation and short path evaporation Artisan systems for the removal of environmental and processing-generated contaminants such as polychlorinated biphenyl (PCBs), dioxins, furans, 3-chloropropane-1,2-diol (3-MCPD), glycidyl esters (GE) from oils such as palm, soybean and marine fats. The oils were also analyzed for free fatty acids, mono and diglycerides, peroxide value, anisidine value, color and flavor. The resulting oils met commercial quality specifications for refined edible oils, with the added advantage of reducing free fatty acids from 3.87 % (for palm oil) to < 0.1 % and removal of over 90 percent of mono and diglycerides, confirming the additional application of Artisan's SPS as a physical refining system and fractionation and recovery of mono and diglycerides. Regulatory specs for PCBs, dioxins and furans, MCPDs, GEs for edible oils were also successfully met.

Introdução: O alecrim, é considerado popularmente como terapêutico, sobre microrganismos de notável importância clínica. A busca de produtos naturais que apresentem uma ação antifúngica eficiente frente a microrganismos resistentes se mostra uma alternativa necessária, assim como em virtude da resistência crescente aos antimicrobianos. Objetivo: Caracterizar a eficiência nas atividades antimicrobianas e antifúngicas do óleo essencial de Alecrim (Rosmarinus officinalis L.). Material e métodos: As buscas foram realizadas em bases de dados bibliográficas — SciELO, BIREME, LILACS e Livro de Fitoterapia. Incluiu-se artigos do período de janeiro de 2006 a dezembro de 2020 com delineamento experimental e observacional, em Inglês, Português e Espanhol. Resultados: As propriedades antimicrobianas do alecrim são atribuídas aos compostos pinenos, 1-8 cineol, borneol, canfeno e cânfora. Estes, geralmente, estão presentes no óleo essencial, o qual demonstrou atividade inibitória sobre as cepas de S. aureus e E. coli, quando utilizado o método de orifício em Ágar, mas não pelo método de difusão em disco, quando avaliadas as culturas de S. aureus, mas apresenta efeito bactericida para a linhagem de S. aureus. Para E. coli, foi verificada sensibilidade a este composto também no método de disco difusão. Frente a Staphylococcus spp, Enterobacter gergoviae, E. amnigenus, Lactobacillus sakei, e L. curvatus, o óleo essencial, apresentou atividade antimicrobiana, caracterizando a sua eficácia perante esses microrganismos. O óleo essencial de R. officinalis L. em relação aos fungos provoca a granulação do citoplasma, desorganização dos conteúdos celulares, rompimento da membrana plasmática e inibição das enzimas fúngicas. A partir do teste de difusão em meio sólido, atividade antifúngica sobre a cepa de C. albicans, foi identificada, assim como atividade antifúngica e antiaderente sobre C. tropicalis. No teste in vitro, mostrou-se eficiência em inibir o crescimento do A. flavus. Conclusão: Através dessa revisão bibliográfica, foi possível identificar a eficiência na atividade antimicrobiana e antifúngica do óleo essencial de Alecrim (Rosmarinus officinalis L.) diante das cepas analisadas. Porém, são necessários estudos sobre as composições e aspectos que podem comprometer a eficácia do óleo essencial, para garantir a segurança e ação do produto.

Introdução: As aves são consideradas dentre todos os grupos animais as mais fáceis de serem observadas na natureza em diversos locais do mundo e seu comportamento social e reprodutivo, que varia entre as espécies, podem ser utilizados como ferramentas para avaliação da estrutura do habitat e sua conservação. Objetivo: O objetivo deste trabalho foi realizar uma revisão bibliográfica com informações acerca do impacto do nidoparasitismo em aves de vida livre, sendo este negativo ou positivo. Material e métodos: Foi realizado um levantamento bibliográfico a partir de 8 artigos entre os anos de 2003 a 2019 com os termos \"nidoparasitismo em aves\", \"parasitas de ninhos\" e \"comportamento reprodutivo das aves\". Os nidoparasitas ou parasitas de ninho são aves que possuem o hábito de não construírem seu próprio ninho, depositando seus ovos em ninhos de outras aves para que estas choquem, criem e alimentem seus filhotes. Enquanto as aves que recebem os filhotes são chamadas de aves hospedeiras. Espécies como Tapera naevia (Saci), Molothrus bonariensis (Chupim), Pintangus sulphuratus (Bem-te-vi), Dromococcyx pavoninus (Peixe-frito-pavonino) e Molothrus oryzivorus (Irauná-grande) possuem este comportamento reprodutivo e quando realizam a postura dos ovos nos ninhos tendem a destruir os ovos das aves hospedeiras causando perdas significativas de algumas espécies como Asthenes luizae (Leinheiro-da-serra-do-Cipó), Xanthopsar flavus (Veste-amarela) e Agelaius cyanopus (Carretão-do-Oeste). Resultados: Segundo um estudo realizado sobre nidoparisitismo em colônias de Cacicus cela (Xexéu), a presença de filhotes nidoparasitas de aves da família Tyrannidae são benéficas contra predadores de ninhos. Enquanto outro estudo sobre filhotes parasitários e hospedeiros em ninhos evidenciou que a taxa de sobrevida de ninhos parasitados é maior quando comparada com ninhos não parasitados, independentemente de serem habitados por filhotes hospedeiros ou parasitas. De acordo com o ecologista Paul Dayton, o nidoparasitismo pode ser utilizado como um método de conservação para espécies de aves que estão ameaçadas pois a maioria dos casos ocorrem em áreas alteradas por atividades humanas. Conclusão: Diante do que foi exposto, é necessário mais estudo sobre este comportamento, visto que é um tema de extrema relevância no auxílio de estudos sobre a ecoevolução em relação ao sistema parasita-hospedeiro.

Introdução: A crescente demanda por produtos naturais tem estimulado pesquisadores a desenvolver processos biotecnológicos para a produção de compostos de aroma, como a fermentação em estado sólido (FES), em que resíduos orgânicos podem ser utilizados como substrato ou suporte inerte. Objetivo: Este trabalho tem como objetivo investigar a produção de compostos de aroma a partir da FES de Geotrichum candidum (CCT 1205) e Trichoderma harzianum (IOC 4042) utilizando borra de café in natura (BC-IN) e desengordurada (BC-DES) como suporte. Material e métodos. Para a fermentação, os suportes sólidos (5 g) foram adicionadas em erlenmeyer (250 ml), e impregnados com uma solução nutritiva contendo glicose, (NH4)2SO4, MgSO4, KH2PO4, KCl, CaCl2.H2O, FeSO4 e ZnSO4. O substrato quanto a solução nutritiva foram autoclavados a 121°C por 15 min e inoculados por 1 mL de suspensão de esporos (1,0 x 106 UFC/mL), e mantidos estacionários a 28°C por 7 dias. Os compostos produzidos foram analisados SPME, CG-EM, e caracterizados com base no índice de similaridade (%), GRAS (generally recognized as safe) e FEMA (Flavor & Extract Manufacturers Association). Resultados: A caracterização química indicou que os suportes BC-IN e BC-DES são compostos de (em % m/m, respectivamente): umidade (8,68 e 8,15), proteínas (12,01 e 11,26), lipídios (10,42 e 2,17) e carboidratos totais (67,30 e 76,40). Vários compostos de aroma foram detectados no extrato do BC-IN e BC-DES fermentado por G. candidum e T. harzianum, provando que a retirada da fração lipídica não demonstrou diferenças significativas nos compostos produzidos. Entre eles, hexanal (95%, GRAS 2557, aroma de fruta), 2-feniletanol (98%, GRAS 2858, aroma de rosa), hexanoato de etila (88%, GRAS 2439, aroma de maçã e abacaxi), e 6-pentil-α-pirona (98%, GRAS 3696, aroma de coco) são particularmente perceptíveis entre os compostos identificados via análise CG-EM. Todos os compostos de aroma identificados neste trabalho são reconhecidos pelo status FEMA e GRAS, dessa forma, o uso deles é seguro. Conclusão: Esses resultados comprovam que a produção de compostos de aroma por G. candidum e T. harzianum através do processo de fermentação da borra de café é viável.

Citrus juice flavor quality traditionally diminishes after thermal processing and continuously during storage. Our prior studies found that four of the five most potent off-aromas formed during orange juice storage had not been identified. The primary emphasis of this project was to characterize and identify those potent flavor degrading aroma volatiles so that methods to control them could be developed and final flavor quality improved. Our original objectives included: 1 Isolate and characterize the most important unidentified aroma impact compounds formed or lost during pasteurization and storage. 2. Determination of thiamine and carotenoid thermal decomposition and Strecker degradation pathways in model solutions as possible precursors for the unidentified off-flavors. 3. Evaluate the effectiveness of an "electronic nose" to differentiate the headspace aromas of from untreated and heat pasteurized orange and grapefruit juices. 4. Use model systems of citrus juices to investigate the three possible precursor pathways (from 2) for flavor impact compounds formed or lost during pasteurization or storage. RESULTS - The components responsible for citrus storage off flavors and their putative precursors have now been identified. Certain carotenoids (b-carotene) can thermally degrade to produce b-ionone and b-damascenone which are floral and tobacco smelling respectively. Our GC-O and sensory experiments indicated that b-damascenone is a potential storage off-flavor in orange juice. Thiamine (Vitamin B1) degradation produces 2-methyl-3-furan thiol, MFT, and its dimer bis(2- methyl-3-furyl) disulfide which both produce meaty, savory aromas. GC-O and sensory studies indicated that MFT is another storage off-flavor. Methional (potato aroma) is another off flavor produced primarily from the reaction of the native amino acid, methionine, and oxidized ascorbic acid (vitamin C). This is a newly discovered pathway for the production of methional and is more dominant in juices than the classic Maillard reaction. These newly identified off flavors diminish the flavor quality of citrus juices as they distort the flavor balance and introduce non-typical aromas to the juice flavor profile. In addition, we have demonstrated that some of the poor flavor quality citrus juice found in the market place is not only from the production of these and other off flavors but also due to the absence of desirable flavor components including several potent aldehydes and a few esters. The absence of these compounds appears to be due to incomplete flavor volatile restoration after the making of juice concentrates. We are the first to demonstrate that not all flavor volatiles are removed along with water in the production of juice concentrate. In the case of grapefruit juice we have documented which flavor volatiles are completely removed, which are partially removed and which actually increase because of the thermal process. Since more that half of all citrus juices is made into concentrate, this information will allow producers to more accurately restore the original flavor components and produce a juice with a more natural flavor. IMPLICATIONS - We have shown that the aroma of citrus juices is controlled by only 1-2% of the total volatiles. The vast majority of other volatiles have little to no direct aroma activity. The critical volatiles have now been identified. The ability to produce high quality citrus juices requires that manufacturers know which chemical components control aroma and flavor. In addition to identifying the critical flavor components (both positive and negative), we have also identified several precursors. The behavior of these key aroma compounds and their precursors during common manufacturing and storage conditions has been documented so manufacturers in Israel and the US can alter production practices to minimize the negative ones and maximize the positive ones.

The specific objectives of approved proposal include to: 1. Elucidate the C6-C2 biochemical pathways leading to the biosynthesis of phenylacetaldehyde, phenylethyl alcohol and phenylethyl acetate in floral tissues of ornamentally important plants, pefunia and roses. 2. Isolate and characterrze genes responsible for the production of these C6-C2 compounds and those involved in the regulation of the pathway using genomic and transcriptomic tools. 3. Determine whether altering the expression of key genes of this pathway can result in changing the aroma characteristics of flowers. Aldehydes are intermediates in a variety of biochemical pathways including those involved in the metabolism of carbohydrates, vitamins, steroids, amino acids, benzylisoquinoline alkaloids, hormones, and lipids. In plants they are also synthesized in response to environmental stresses such as salinity, cold, and heat shock or as flavors and aromas in fruits and flowers. Phenylacetaldehyde along with 2-phenylethanol and its acetate ester, are important scent compounds in numerous flowers, including petunias and roses. However, little is known about the biosynthesis of these volatile compounds in plants. We have shown that the formation PHA and 2-phenylethanol from Phe does not occur via trans-cinnamic acid and instead competes with the key enzyme of phenypropanoid metabolism Pheammonia-lyase (PAL) for Phe utilization. Using functional genomic approach and comparative gene expression profiling, we have isolated and characterized a novel enzyme from petunia and rose flowers that catalyzes the formation of the Ca-Czcompound phenylacetaldehyde (PHA) from L-phenylalanine (Phe) by the removal of both the carboxyl and amino groups. This enzyme, designated as phenylacetaldehyde synthases (PAAS), is a bifunctional enzyme that catalyzes the unprecedented efficient coupling of phenylalanine decarboxylation to oxidation, generating phenylacetaldehyde, CO2, ammonia, and hydrogen peroxide in stoichiometric amounts. Down-regulation of PAAS expression via RNA interference-based (RNAi) technology in petunia resulted in no PHA emission when compared with controls. These plants also produced no 2-phenylethanol, supporting our conclusion that PHA is a precursor of 2-phenylethanol. To understand the regulation of scent formation in plants we have also generated transgenic petunia and tobacco plants expressing the rose alcohol acetyltransferase (RhAAT) gene under the control of a CaMV-35S promoter. Although the preferred substrate of RhAAT in vitro is geraniol, in transgenic petunia flowers, it used phenylethyl alcohol and benzyl alcohol to produce the corresponding acetate esters, not generated by control flowers. These results strongly point to the dependence of volatile production on substrate availability. Analysis of the diurnal regulation of scent production in rose flowers revealed that although the daily emission of most scent compounds is synchronized, various independently evolved mechanisms control the production, accumulation and release of different volatiles. This research resulted in a fundamental discovery of biochemical pathway, enzymes and genes involved in biosynthesis of C6-C2s compounds, and provided the knowledge for future engineering plants for improved scent quality.