β-Glucans from Yeast—Immunomodulators from Novel Waste Resources

Round 1

Reviewer 1 Report

I found the review paper to be overall well written dealing with β-glucans which have been reported as effective in lowering D-dimer levels, one of the most common side effect of COVID-19. However, I have some suggestions which could improve the quality of the proposed paper.  In the section 2. Yeast as source of β-glucans, recent study on valorization of winery spent yeast waste could be discussed. In section 9. Extraction and Characterization, please elucidate abbrevation T.C.A.  Since ultrasound assisted extraction (UAE) is mentioned as a potential extraction technique it would be useful to cite optimal parameters in UAE for maximization amount of extracted β-glucans. In addition, microwave assisted extraction could be also added alone or as pretreatment prior to UAE for extraction of ß-glucans.   

Author Response

I found the review paper to be overall well written dealing with β-glucans which have been reported as effective in lowering D-dimer levels, one of the most common side effect of COVID-19. However, I have some suggestions which could improve the quality of the proposed paper. 

Thank you for the synopsis of our manuscript and for the words of praise. We have addressed your suggestions below.

Suggestion 1:

In the section 2. Yeast as source of β-glucans, recent study on the valorization of winery spent yeast waste could be discussed.

Thank you, this is a very relevant study for this review. We have included this into Section 3- Production of Yeast b-glucans from Waste Streams; Page;3 Lines;142-152 . The section added- “A novel source for commercial synthesis of yeast b-glucans was investigated by Varelas et al. The group isolated b-glucans for the first time from winery spent yeast biomass. During the winemaking process, a byproduct known as wine lees is produced. Most byproducts include spent yeasts, bacteria, tartaric acid, ethanol, phenolics and pigments. Thus, b-glucans can be sourced from yeast waste biomass that accumulates in wine tanks throughout the winemaking process. This study showed that the b-glucans isolated contained some amount of tartaric acid and polyphenols, which could not be omitted. Considering wine lees, especially red ones, are more complex mixes than brewery wastes, the purity of b-glucan in wine lees samples is lower than the purity reported by other studies for brewery wastes (Varelas et al., 2016). Nonetheless, this work identifies a valuable waste source of b-glucans that are most often disposed of in landfills.”

Suggestion 2:

In section 9. Extraction and Characterization, please elucidate abbrevation T.C.A. 

Thank you, this has been amended to Trichloroacetic acid (T.C.A)

Suggestion 3:

Since ultrasound assisted extraction (UAE) is mentioned as a potential extraction technique it would be useful to cite optimal parameters in UAE for maximization amount of extracted β-glucans.

Thank you, this is method is relevant to discuss as it is a green technique and we are highlighting waste valorisation so it all ties well. We have included this into Section 9 Extraction and Characterisation of b-glucans; Page;12  Lines; 517-556.  The section added “Ultrasound-assisted extraction (UAE) is a green extraction technique that avoids using strong solvents (Chémat & Strube, 2015). UAE accelerates the extraction of molecules but also preserves the structural and molecular characteristics (Flórez-Fernández & González Muñoz, 2017).

Ultrasounds (US) are mechanical waves having a frequency over 20 kHz. Two types of US exist low-intensity and high-intensity US. The low-intensity US are used to test physicochemical properties in the food industry. For the extraction of chemicals of interest, high-intensity ultrasounds are used at frequencies of 20 - 100 kHz (Flórez-Fernández & González Muñoz, 2017).

The effect of operational variables on the UAE b-glucans from barley was investigated in a study by Benito-Román et al. The operational variables used in this investigation were amplitude, time, and cycles in conjunction with varied ultrasonic powers to increase extraction performance and molecular weight in beta-glucan extraction. The extraction performance is closely related to amplitude and time. When compared to conventional extraction, UAE reduces extraction times and energy usage. The data shows that extraction yield relies on amplitude and time, whereas molecular weight decreases with time. The highest extraction yield (66%) is obtained by supplying the most energy (962.5 kJ/L), resulting in the lowest molecular weight (269 kDa). Reduced treatment intensity (energy output of 170 kJ/L) reduces extraction yield to 444.3% but raises molecular weight to 461 kDa. However, while the UAE process improves the extraction yield and molecular weight of b-glucans over stirred tank extraction (3 h, 55 °C, 1000 rpm), the main effect of ultrasounds is the reduction of process time and energy consumption (3 min versus 3 h and 170 kJ/L versus 1460 kJ/L) (Benito-Román et al., 2013). 

Another similar study determined parameters of UAE of polysaccharides from white button mushroom (A. bisporus). The efficient extraction parameters were 230 W ultrasonic power, 70°C extraction temperature, 62 min extraction duration, and 30 ml/g W/M ratio, yielding 6.02 per cent A. bisporus polysaccharides (ABPS). ABPS had a molecular weight of 158 kDa (Tian et al., 2012)’’.

 

Suggestion 4:

In addition, microwave assisted extraction could be also added alone or as pre-treatment prior to UAE for extraction of ß-glucans.   

Thank you, this again is an important extraction method that is very relevant to this study.

We have included this into Section 9 Extraction and Characterisation of b-glucans; Page;13 Lines; 543-556.

The section added “Additionally, combining ultrasonic waves with microwave-assisted extraction (MAE) for the separation of molecules of interest might yield even more significant advantages (Flórez-Fernández & González Muñoz, 2017). MAE uses microwave energy to heat the extraction solvent directly. The increased pressure and temperature that microwave extraction provides keep the water in a liquid state and above the boiling point. In addition to reduced CO2 emissions and reduced usage of polluting solvents, this technique is deemed environmentally friendly because of its improved efficiency and lower energy use. In addition to being more effective and selective, this non-contact heat source may also speed up energy transfer, start-up, reaction to heating control, and eliminate temperature gradients (Smiderle et al., 2017).

Studies have shown that the highest yield of b-glucans from mushrooms using MAE was obtained at 180 °C and 30 min (Gil-Ramírez et al., 2019).   Other studies have demonstrated that MAE had high reproducibility and was efficient and fast at b-glucan extraction from mushrooms (Smiderle et al., 2017). MAE can be used alone as an extraction technique or combined with other methods like UAE.

Reviewer 2 Report

- Few linguistic errors need to be corrected.

- Many references are from before 2013. Please consider updating them if possible

Specific comments and suggestions for authors are reported in the attached file.

Comments for author File: Comments.pdf

Author Response

Thank you for your comments, we have attached our responses in a PDF.

Author Response File: Author Response.pdf