Effect of the combined application of heat treatment and proteases on protein stability and volatile composition of Greek white wines

Abstract

Aim: This work evaluates the effects of the combined use of heat treatment (HT, 75 °C, 2 min) and proteases (P) on the protein stability and volatile composition of two white wines, obtained from the Greek cv. Assyrtiko and Moschofilero. Methods and results: Wine protein stabilization was assessed by heat test, using RP-HPLC determination of pathogenesis-related proteins (PRP) and by sodium dodecyl-sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The impact of bentonite and P + HT treatment on wine aroma profile was evaluated by GC-MS with liquid-liquid extraction. According to the heat test, in Assyrtiko wine the level of stability achieved with P + HT was comparable with that obtained by bentonite fining and for Moschofilero wine -where protein instability was higher -bentonite was more efficient. RP-HPLC profiles showed that, in general, higher percentages of chitinases (CH) than thaumatin-like proteins (TLP) were removed by both bentonite and P + HT, with a similar efficiency for the two treatments and sometimes better performances for the latter. Conversely, TLP were removed more efficiently by bentonite, even if some fractions were eliminated to a slightly higher extent by proteases. In the conditions of the experiment, bentonite resulted in minor changes to the wine aroma profile. However, heating during protease treatment modified wine volatile composition, reducing the concentration of esters produced during fermentation while simultaneously increasing the contents of certain esters characteristic of aging such as ethyl lactate. Conclusions: The combination of proteases and heat treatment may be a promising technique for protein stabilization of wines. However, further investigations are needed to optimize the time:temperature ratio of the heat treatment in order to obtain the maximum protease activity and the minimum thermal deterioration of the wine quality. Significance and impact of the study: The results of this study have a practical interest for both the scientific community and wine sector, contributing to knowledge of the efficacy and limitations of the use of protease enzymes for wine stabilization.Assyrtiko, bentonite fining, Moschofilero, protease, protein stability, wine aroma

Advances in White Wine Protein Stabilization Technologies

“…The table includes the operating conditions, range of protein reduction (%), and time of application for each technology. AF 2-3 g/L 67-95% [24][25][26] Physical-enzymaticmixed treatments High power ultrasound AF 30%/10 min; 60-90%/5-10 min n/a [27] Heat + enzymes BF 75 • C for 1 min + 15 mg/L of enzymes 1 81-84% [5] AF 75 • C for 2 min + 2 mL/L of enzymes 2 80-90% for CHI [28] Ultrafiltration AF 80% permeate/20% retentate, 10 kDa membrane n/a [29] Ultrafiltration + heat + enzymes AF 62 • C for 10 min + 30 mg/L of enzymes 1 30-96% [29] Immobilized enzyme supported on chitosan AF Continuous PBR 3 : 0.3-15 mL/min flow and 106-260 g/L 4-68% 61-63% [30,31] Adsorption-based treatments Magnetic nanoparticles coated with acrylic acid AF 10 W by 10 min (plasma deposition) and 13.3-25 g/L for 10 min >90% [32][33][34] Zeolites AF 4-8 g/L of zeolites for 1-3 h >90% [35,36] Roasted grape seeds powder BF 5-15 g/L for 1 h 37-85% [37,38] AF 25-32 g/L for 1 h 90-98% [37] Zirconium DF 25 g/L pellet in metallic cage for 3 days ~90% [39] AF Batch: 25 g/L for 72-192 h >70% [22] Continuous: 175-300 BV 4 (~5.7-3.3 g/L) for 30 min of residence time ~42% [40] Closed-loop operation: pellet packed (6.5 L), 300 L/h flow rate for 8-139 h ~54-60% [41,42] 1 Enzyme type: Proctase, prepared containing aspergillopesin I and II. 2 Enzyme type: Two proteases in aspergillopepsin-containing liquid preparations obtained from Aspergillus niger.…”

“…Contrary to Marangon et al [5], who performed the treatments before fermentation, Comuzzo et al and Sui et al [28,29] carried out the experiments after fermentation (Figure 2). The treatment used by Comuzzo et al [28] (Figure 2a) consisted of heating the wine for 2 min at 75 °C plus the addition of enzymes (1-2 mL/L). Comuzzo et al [28] studied the effect on protein stability and volatile compound losses of AGP + heat treatment in two varieties of Greek white wines (Moschofilero and Assyrtiko).…”

“…Mixed treatments of heating plus enzyme addition have better results than separate treatments. The heat treatment causes the proteins to unfold (55 • C for CHIs and at 62 • C for TLPs) [6], which in turn, makes the proteins easier to break down by the active aspergillopepsin enzymes (enzymatic activation used in heat treatments is between 60 and 80 • C from 1 to 10 min) [5,28,29]. However, Falconer et al [6] noticed that some TLPs could be refolded, unlike CHIs, which unfold irreversibly.…”

“…Against scientific evidence and despite regulatory approval in the European Union (2010), Australia (2014), New Zealand (2014), and associated export markets [5], there are concerns about the use of this technology; hence, this method is not widely used [29]. Contrary to Marangon et al [5], who performed the treatments before fermentation, Comuzzo et al and Sui et al [28,29] carried out the experiments after fermentation (Figure 2). The treatment used by Comuzzo et al [28] (Figure 2a) consisted of heating the wine for 2 min at 75 °C plus the addition of enzymes (1-2 mL/L).…”

“…The treatment used by Comuzzo et al [28] (Figure 2a) consisted of heating the wine for 2 min at 75 °C plus the addition of enzymes (1-2 mL/L). Comuzzo et al [28] studied the effect on protein stability and volatile compound losses of AGP + heat treatment in two varieties of Greek white wines (Moschofilero and Assyrtiko). They observed that this Contrary to Marangon et al [5], who performed the treatments before fermentation, Comuzzo et al and Sui et al [28,29] carried out the experiments after fermentation (Figure 2).…”