Production of medium-chain volatile flavour esters in Pichia pastoris whole-cell biocatalysts with extracellular expression of Saccharomyces cerevisiae acyl-CoA:ethanol O-acyltransferase Eht1 or Eeb1

Medium-chain volatile flavour esters are important molecules since they have extensive applications in food, fragrance, cosmetic, paint and coating industries, which determine different characteristics of aroma or taste in commercial products. Biosynthesis of these compounds by alcoholysis is catalyzed by acyl-CoA:ethanol O-acyltransferases Eht1 or Eeb1 in Saccharomyces cerevisiae. In this study, these two yeast enzymes were selected to explore their preparations as the form of whole cell biocatalysts for the production of volatile flavour esters. Here, the novel whole cell biocatalysts Pichia pastoris yeasts with functional extracellular expression of Eht1 or Eeb1 were constructed. Flavour production was established through an integrated process with coupled enzyme formation and ester biosynthesis in the recombinant yeasts in one pot, leading to the formation of volatile C6–C14 methyl and ethyl esters from wort medium. Interestingly, there is no significant difference between P. pastoris-EHT1 and P. pastoris-EEB1 in substrate preference during flavour biosynthesis, indicating a similar role of Eht1 and Eeb1 in P. pastoris cells, in contradiction with previous findings in S. cerevisiae to some extent. Consequently the study not only provides a greater understanding of these two enzymes in a heterogeneous host, but also demonstrated the positive effect of the recombinant Eht1 and Eeb1 in ester formation by P. pastoris live cells, potentially paving the way towards achieving efficient production of volatile flavour by an integrated biocatalytic system composed of recombinant enzyme production and flavour biosynthesis

Changes in the aroma profile of three Vitis vinífera cultivars - Gran Negro, Mouratón and Brancellao - depending on ripening and the position of the berries into the bunch

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Skin and flesh odorants - free and bound form - in three red grapes depending on harvesting time and berry position into the bunch

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Aroma potential of Brancellao grapes from different cluster positions

In this study the presence of aroma compounds in grapes of Brancellao (Vitis vinifera L.) was investigated in order to obtain its aroma potential fingerprint. It is well known that differences exist in aromatic compounds amongst grapevine varieties at ripening stages. Within the framework of an increasingly competitive market, the chance of obtaining different wines from vines of the same variety grown at the same vineyard is becoming of increasing importance. This can be done through the managing of the vineyard, but also some wineries have assayed the separation of the tip and shoulder berries of the clusters of a specific variety with this objective. In this work it is evaluated that, in the final stages of maturation, differences exist in the probable alcoholic degree, total acidity of the must, as well as in the aromatic composition of skin and flesh of berries coming from the tips and shoulders of the clusters. Gas chromatography coupled to mass spectrometry (GC-MS) was used to determine the aromatic composition, in the skin and flesh of each sample, either tip or shoulder berries from the clusters. The obtained results showed that there was not variability for the probable alcoholic degree and total acidity between the shoulders and tips, whereas there was variability for their aromatic composition. For the berries from the tips of the clusters most of volatiles were found in the flesh (except aldehydes) and spicy and floral nuances (with the only exception of β-ionone) were in higher proportions. For the berries from the shoulders of the clusters, most of volatiles were found in the skin (monoterpenes, norisoprenoids, aldehydes, and C6 alcohols), where the flesh was slightly richer in aromatic alcohols, volatile phenols and pantolactone; β-ionone and herbaceous nuances were in higher proportions. These results are promising for those wineries that are considering the chance of separating berries from tips and shoulders of the clusters for the elaboration of different quality wines. © 2011 Elsevier Ltd. All rights reserved.This work was Granted by EU FEDER funds, by Ministry of Science and Technology (Spain) INIA RF-2008-00002-C02 and by the INCITE09PXIB383322 PR (Autonomous Community Government in N.W. Spain) contractsPeer Reviewe

Active odorants in Mouratón grapes from shoulders and tips into the bunch

The opportunity for obtaining different wines from the same variety cultivated in the same vineyard is becoming of increasing importance. This is why some wine cellars have started to assay the separation of the tips and shoulders berries of the clusters of a specific variety with this objective. In this work, the study is focused on berries of Mouratón (Vitis vinifera L.) to investigate if, in the latter stages of ripening, differences exist in the probable alcoholic degree, total acidity of the must, as well as in the aromatic composition of skin and flesh of berries coming from the tips and shoulders of the clusters. Gas chromatography coupled to mass spectrometry (GC-MS) was used to determine the aromatic composition. The obtained results showed that there was not variability for the probable alcoholic degree and total acidity between the tips and shoulders, whereas there was variability for their aromatic composition during ripening. These results are promising for those wine cellars that are considering the separation of berries from tips and shoulders of the clusters for the elaboration of different quality wines. © 2012 Elsevier Ltd. All rights reserved.EU-FEDER; Ministry of Science and Technology (Spain) INIA RF-2008-00002-C02; INCITE09 383 322 PR (Xunta de Galicia)Peer Reviewe