In recent years, cold temperature technology has gained growing relevance in modern winemaking due to its positive impact on the aromatic profile and sensory attributes of wines – improving the overall wine quality in terms of its colour, aroma and final taste. This technology can be strategically implemented at different stages of the enological process to optimize both chemical and sensory attributes¹. Additionally, cold temperatures in the form of cryopreservation can be used to preserve communities of microorganisms for long periods of time. Within the Eco2Wine project, we aim to preserve the microbial consortia as untouched as possible as a “time-capsule” of biodiversity for future use of winemakers.

In this article, we will explore why cryopreservation is necessary for the preservation of wine microbial consortia, as well as the benefits of using cold temperature technology during maceration and fermentation to improve wine quality.

Cryopreservation 

The use of low-temperature technologies in oenology extends beyond the processing of grapes to cryopreservation, a biological application using ultra-low temperature (below 0℃) for the long-term preservation of microorganisms.

At ultra-low temperatures, microorganisms are mostly inactivated from evolution, hence maintaining their genetic stability. Although sometimes chemical changes within cells could happen due to free radical formation and ionizing radiation¹, cryopreservation is still considered a good option for long term preservation. It does not involve routine manipulation which reduces the risk of contamination and human errors.

Why is cryopreservation necessary in oenology? 

In oenology, the concept of terroir refers to the combination of environmental factors contributing to the wine quality. These environmental factors shape the microbial diversity present in the vineyard². This being said, each vineyard has its own unique microbial consortia that contribute to the unique taste and aroma of wine produced in the region. Thus, cryopreservation becomes necessary with the increasing interest in indigenous yeast strains and their application to small and specific wine production.

Problems in preservation of indigenous strains 

However, the preservation of indigenous yeasts presents some problems. First, only a small percentage of yeasts can be actively dried. In the past, yeasts were preserved with traditional methods, using agar and liquid culture; it only preserved less than 1% of the total microbial population³. Furthermore, some yeasts exist in a viable but non-culturable state (VBNC); a phenomenon where the yeasts are alive but unable to grow and form colonies on media⁴. With the traditional methods, they are unable to be isolated and stored.

Secondly, there is an increasing interest in preserving microbial consortia – a community of two or more microorganisms. The cryopreservation of microbial consortia is not straightforward – it requires time and research for the protocol optimization. Different yeast species will have different biological responses to the cryopreservation condition⁵, making the optimization process complex.

Optimization of cryopreservation protocol

Today, the optimal cryopreservation protocol for winemaking yeasts is still not well-defined yet, especially for microbial consortia. One of the research aims of Eco2Wine is to come up with a protocol that ensures the highest quality of cryopreserved indigenous yeasts. This will support the overall project goal of developing techniques for more natural wine production.

Impact on grape aroma compounds and sensory properties of wine

Cold temperature technology can also be applied during maceration and fermentation to improve the wine quality. ​Studies have shown that the use of cold temperature technology during maceration enhances the aromatic profile of wines^6,7. These methods involve freezing grape berries to facilitate the extraction of aroma compounds, leading to wines with more intense and stable aromas. Freezing grapes before winemaking significantly affects their chemical composition. Research conducted by Carillo et al.⁸ found that phenolic acids (such as gallic and caffeic acid) and polyphenols differ between wines made from fresh and frozen grapes.

Similarly, freezing also impacts acetaldehyde levels and volatile compounds, with frozen grape juice having a different aroma profile compared to fresh juice⁹. The effects of freezing vary depending on the grape variety and the specific freezing method used. While fresh juice is preferable for analyzing volatile compounds, long-term storage at -80°C or in liquid nitrogen is recommended to minimize biochemical changes¹⁰. Additionally, freezing reduces anthocyanins, total phenolics, and terpenic alcohols, which are important for wine color and aroma⁸.

Apart from that, wines made from grapes treated with liquid CO₂ (inertized wine) had better color and phenol concentration compared to untreated wines. Additionally, non-trained judges preferred these wines, suggesting they align with consumer tastes⁸. Cold pre-fermentation maceration also enhances aroma compounds (terpenes, thiols, esters, and phenols) by increasing their extraction from the grape skin. This process can modify the nutrient composition of the grape must, leading to wines with improved aromatic complexity and overall quality¹¹. Overall, research suggests that the application of cold temperature technology enhances aroma intensity, making them a valuable tool for improving wine quality¹².