Yeast and its ability to release thiols

by | Aug 1, 2020 | Oenology research, Winetech Technical

PHOTO: Shutterstock.

Yeast strains vary significantly in their ability to produce varietal thiols.

Introduction

The varietal thiols, mainly 3-mercaptohexan-1-ol (3MH), 3-mercaptohexyl acetate (3MHA) and 4-mercapto-4-methyl-pentan-2-one (4MMP) are the three major varietal thiols contributing to tropical fruit aroma wines. These aromatic compounds are considered to be impact compounds for Sauvignon blanc wines, however, elevated levels of thiols have also been found in South African Chenin blanc and Colombar wines.

Thiol precursors and yeast

The varietal thiols, 3MH and 4MMP are present in the non-aromatic form in the juice and are formed and/or released from precursors during fermentation. Yeast takes up conjugated precursors from grape juice and then cleaves the precursor, releasing the corresponding free thiol. Non-odourant cysteinylated and glutathionylated precursors, such as Cys-3MH, Cys-Gly-3MH, GSH-3MH, Cys-4MMP, Cys-Gly-4MMP and GSH-4MMP, have been identified as conjugated precursors in the grape must.1-4 However, studies have also shown that these precursors are not the exclusive source of varietal thiols and other mechanisms and unidentified precursors also contribute significantly to the final content of varietal thiols in wine.5,6 Unlike 3MH and 4MMP, 3MHA does not have a conjugated precursor in the must and is formed during the esterification of 3MH.

Even though the exact mechanisms for the release and/or formation of these odoriferous thiols are still not entirely elucidated, it is clear that the yeast strain used to perform the fermentation is one of the most important factors affecting the thiol production and the thiol concentration in the final wine.7 Yeast strains differ significantly in the capability to produce thiols. This ability (or lack thereof) can be exploited by the winemaker to specifically craft the desired wine style by selecting a yeast strain with specific properties and capabilities.

It’s all in the genes

Before thiol liberation, the yeast needs to transport the conjugated precursor from the juice into the yeast cell. For this transportation, the yeast needs to be in possession of general amino acid transporters located in the cell membrane. Once in the cell, the glutathionylated precursor is first transformed into the cysteinylated precursor before the aromatic thiol can be released. This transformation entails a complex mechanism involving multiple genes.8

For the cleavage of the cysteinylated precursors (and subsequent release of the aromatic thiols), a specific enzyme, namely the carbon-sulphur β-lyase enzyme, is required. Various genes have been identified as being responsible for the yeasts’ production capabilities of cleaving enzymes including BNA3, CYS3, STR3 and GLO1. However, the main gene responsible for the release of thiols is the IRC7 gene. Studies done in a synthetic wine medium with precursors present have shown that a full-length copy of IRC7 is required for the cleavage of precursors and the deletion of the gene resulted in most yeast strains being incapable of converting the conjugated precursors into the aromatic thiols.9

This gene could potentially serve as a molecular marker to predict a yeast’s potential to release thiols. That said, in a juice medium (and not a synthetic medium with limited precursors available), the presence of the IRC7 gene was not a prerequisite for the production of varietal thiols,10 supporting the hypothesis that other unknown precursors also contribute significantly to the final concentration of varietal thiols in wine.

The esterification of 3MH to form 3MHA also requires a gene encoding alcohol acetyltransferase (ATF1).11

Yeast strain variability

Several studies have reported the limited capacity of most Saccharomyces cerevisiae yeast strains to release varietal thiols from the corresponding non-volatile precursors (usually less than 5% converted).12

A study13 assessed the ability of 82 different yeast strains (not all commercially available) to produce thiols from a grape-like medium containing realistic concentrations of conjugated thiol precursors. Results showed a 20-fold difference between the yeasts’ ability to release 3MH and a 35 fold difference in the ability to produce 4MMP. The yeasts were categorised according to the amount of thiols produced during fermentation and results showed that the majority (70%) of the strains tested were considered to be low releasers of thiols, 20% were moderate releasers and 10% were considered high releasers.

In most cases, the low releasers possessed an inactive IRC7 gene, which limited the ability to release thiols, while the moderate and high releasers had an active or partially active IRC7 gene.

Among the selection of commercial yeast strains tested, the strains recommended by suppliers for the production of white wine produced relatively higher concentrations of volatile thiols when compared to strains that were recommended for the production of red wine only or the production of both red and white wine. This shows some active selection from yeast manufacturers for strains with a higher thiol-releasing capacity for the production of fruit-driven white wines.

Non-conventional yeasts

The use of non-conventional yeasts can be an important tool for thiol production during fermentation. Certain species, such as Torulaspora delbrueckii, Kluyveromyces marxianus and Metschnikowia pulcherrima, show significant potential with marked β-lyase activity and thiol production, but with high strain dependency.14,15 Mixed fermentation of Sauvignon blanc must with Pichia kluyveri and Saccharomyces cerevisiae also showed a marked increase in 3MHA content,16 while sequential fermentation with Torulaspora delbrueckii and Saccharomyces cerevisiae showed a significant increase in 4MMP production compared to fermentation with Saccharomyces cerevisiae alone.17

As multi-starter fermentations are increasingly being used for organoleptic and quality improvement, the possible advantages regarding thiol production should also be considered.

Conclusions

Commercial yeast manufacturers offer a wide range of yeast strains often with recommendations based on the ability to produce certain attributes. Knowledge of yeast genes required provides tools to distinguish potential yeast strains based on the ability to produce and convert thiols. This will help the winemaker make an informed decision when selecting a yeast strain based on specific target market segments.

Abstract

Studies have shown specific genes are involved in the process of releasing aromatic thiols from the conjugated precursors and could serve as markers to identify yeast strains with high thiol releasing potential. A better understanding of the genes and mechanisms involved will allow winemakers to more reliably produce wines in their desired wine style.

References

  1. Tominaga, T., Peyrot des Gachons, C. & Dubourdieu, D., 1998. A New Type of Flavour Precursors in Vitis Vinifera L. Cv. Sauvignon Blanc: S-Cysteine Conjugates. J. Agric. Food Chem. 46, 5215 – 5219.
  2. Tominaga, T., Masneuf, I. & Dubourdieu, D., 1995. A S-Cysteine Conjugate, Precursor of Aroma of White Sauvignon. J. Int. Sci. Vigne Vin 29(4), 227 – 232.
  3. Des Gachons, C.P., Tominaga, T. & Dubourdieu, D., 2002. Sulfur Aroma Precursor Present in S-Glutathione Conjugate Form: Identification of S-3-(Hexan-1-Ol)-Glutathione in Must from Vitis Vinifera L. Cv. Sauvignon Blanc. J. Agric. Food Chem. 50(14), 4076 – 4079. https://doi.org/10.1021/jf020002y.
  4. Fedrizzi, B., Pardon, K.H., Sefton, M.A., Elsey, G.M. & Jeffery, D.W., 2009. First Identification of 4-S-Glutathionyl-4-Methylpentan-2-One, a Potential Precursor of 4-Mercapto-4-Methylpentan-2-One, in Sauvignon Blanc Juice. J. Agric. Food Chem. 57(3), 991 – 995.
  5. Pinu, F.R., Jouanneau, S., Nicolau, L., Gardner, R.C. & Villas-Boas, S.G., 2012. Concentrations of the Volatile Thiol 3-Mercaptohexanol in Sauvignon Blanc Wines: No Correlation with Juice Precursors. Am. J. Enol. Vitic. 63(3), 407 – 412. https://doi.org/10.5344/ajev.2012.11126.
  6. Harsch, M.J., Benkwitz, F., Frost, A., Colonna-Ceccaldi, B., Gardner, R.C. & Salmon, J.M., 2013. New Precursor of 3-Mercaptohexan-1-Ol in Grape Juice: Thiol-Forming Potential and Kinetics during Early Stages of Must Fermentation. J. Agric. Food Chem. 61(15), 3703 – 3713. https://doi.org/10.1021/jf3048753.
  7. Cordente, A.G., Curtin, C.D., Varela, C. & Pretorius, I.S., 2012. Flavour-Active Wine Yeasts. Appl. Microbiol. Biotechnol. 96(3), 601 – 618. https://doi.org/10.1007/s00253-012-4370-z.
  8. Belda, I., Ruiz, J., Esteban-Fernández, A., Navascués, E., Marquina, D., Santos, A. & Moreno-Arribas, M.V., 2017. Microbial Contribution to Wine Aroma and Its Intended Use for Wine Quality Improvement. Molecules 22(2). https://doi.org/10.3390/molecules22020189.
  9. Santiago, M. & Gardner, R.C., 2015. Yeast Genes Required for Conversion of Grape Precursors to Varietal Thiols in Wine. FEMS Yeast Res. 15(5), 1 – 10. https://doi.org/10.1093/femsyr/fov034.
  10. Roncoroni, M., Santiago, M., Hooks, D.O., Moroney, S., Harsch, M.J., Lee, S.A., Richards, K.D., Nicolau, L. & Gardner, R.C., 2011. The Yeast IRC7 Gene Encodes a β-Lyase Responsible for Production of the Varietal Thiol 4-Mercapto-4-Methylpentan-2-One in Wine. Food Microbiol. 28(5), 926 – 935. https://doi.org/10.1016/j.fm.2011.01.002.
  11. Swiegers, J.H., Bartowsky, E.J., Henschke, P.A. & Pretorius, I.S., 2005. Yeast and Bacterial Modulation of Wine Aroma and Flavour. In Australian Journal of Grape and Wine Research; Blair Francis, M.E., Pretorius, I.S., R. J., Ed.; The Australian Wine Research Institute: Glen Osmond, Australia; Vol. 11, pp 139 – 173. https://doi.org/10.1111/j.1755-0238.2005.tb00285.x.
  12. Ruiz, J., Kiene, F., Belda, I., Fracassetti, D., Marquina, D., Navascués, E., Calderón, F., Benito, A., Rauhut, D. & Santos, A., et al., 2019. Effects on Varietal Aromas during Wine Making: A Review of the Impact of Varietal Aromas on the Flavor of Wine. Appl. Microbiol. Biotechnol. 103(18), 7425 – 7450. https://doi.org/10.1007/s00253-019-10008-9.
  13. Cordente, T., Schmidt, S. & Curtin, C., 2017. Understanding Differences among Wine Yeast Strains in Their Ability to Release “tropical” Thiols. AWRI Tech. Rev. 228, 6 – 10.
  14. Belda, I., Ruiz, J., Navascués, E., Marquina, D. & Santos, A., 2016. Improvement of Aromatic Thiol Release through the Selection of Yeasts with Increased β-Lyase Activity. Int. J. Food Microbiol. 225 (March), 1 – 8. https://doi.org/10.1016/j.ijfoodmicro.2016.03.001.
  15. Zott, K., Thibon, C., Bely, M., Lonvaud-Funel, A., Dubourdieu, D. & Masneuf-Pomarede, I., 2011. The Grape Must Non-Saccharomyces Microbial Community: Impact on Volatile Thiol Release. Int. J. Food Microbiol. 151(2), 210 – 215. https://doi.org/10.1016/j.ijfoodmicro.2011.08.026.
  16. Anfang, N., Brajkovich, M. & Goddard, M.R., 2009. Co-Fermentation with Pichia Kluyveri Increases Varietal Thiol Concentrations in Sauvignon Blanc. Aust. J. Grape Wine Res. 15(1), 1 – 8. https://doi.org/10.1111/j.1755-0238.2008.00031.x.
  17. Belda, I., Ruiz, J., Beisert, B., Navascués, E., Marquina, D., Calderón, F., Rauhut, D., Benito, S. & Santos, A., 2017. Influence of Torulaspora Delbrueckii in Varietal Thiol (3-SH and 4-MSP) Release in Wine Sequential Fermentations. Int. J. Food Microbiol. 257 (June), 183 – 191. https://doi.org/10.1016/j.ijfoodmicro.2017.06.028.

– For more information, contact Carien Coetzee at carien@basicwine.co.za.

 

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