Recovery of Residual Brewer’s Yeast by Electroactivation

Yeasts resulting from the brewing process (RBY) are a valuable by-product, with an important content of minerals, vitamins and, especially, proteins. The purpose of the research was the electroactivation of RBY and the simultane-ous obtaining of two products—protein concentrates and hydrolyzed protein from residual brewer’s yeast. Electroactivation is a non-residual process, without the use of chemical reagents and relatively inexpensive. The variation of the electroactivation conditions allowed the separation of 90% - 94% of the proteins in the form of protein concentrates. During the process, it is attested to increase the pH value and decrease the redox potential, which characterizes the multiple redox processes that take place in the cathode cell, including sedimentation at the isoelectric point. The presence of albumin in the protein fractions of RBY allows the formation of protein complexes with calcium, attributing a higher biological value to the obtained products.


Introduction
One of the objectives of the circular economy is the efficient recovery of agroindustrial waste [1]. The coherent application of separation processes for the recovery of value-added compounds, such as bioactive peptides from agro-industrial and biotechnological by-products depends on the development of integrated processes adapted to the specificity of these materials. Brewery residues are a major problem in the brewing business, due to the large volumes generated daily [2]. The holistic analysis of the exploitation of all residues in the beer industry is How to cite this paper: Tarna, R., Vrabie, E., Paladii, I. and Sturza, R. (2021) Recovery of Residual Brewer's Yeast by Electroactivation. Food and Nutrition Sciences, amino acid profiles of the extracts, compared to Saccharomyces cerevisiae and rootlets, another by-product of the beer industry, RBY has a high level of essential amino acids, as shown in Figure 1 [17] [18] [19].
RBY processing encompasses a wide range of methods or a combination of several processes. Enzymatic hydrolysis has been used successfully in the production of peptides and protein hydrolysates from RBY. The hydrolysates obtained require further efficient processing, namely the use of membranes, which is an    Extraction of peptides in protein hydrolysates from RBY is of particular interest in non-residual processing. The properties of peptides and proteins depend on their profile and structure. Thus, their separation from mixtures with complex composition must be achieved by light methods (low temperatures and neutral pH values) with high selectivity, in order to maintain the structural and physico-chemical characteristics of the molecules [24] [25] [26]. Separation performance is determined by membrane selectivity and permeate flow, which are dependent on operating conditions (temperature, pressure, process configuration, module characteristics, cleaning procedure), membrane properties (membrane material and structure, pore size) and feed characteristics (pH, concentration, composition and physico-chemical characteristics) [27]. The objective of selective separation is also an important aspect of the design of the fractionation process [20].
In the context of protein hydrolysate extraction, membrane processes have the ability to maintain protein stability throughout the process and allow for highefficiency separation, which is possible at low temperatures. They do not require the use of solvents and other chemicals [28]. Several biotechnological and phar-  [29]. RBY has a high value of chemical oxygen use (COD) of 0.53 kg/hL, it cannot be eliminated in wastewater streams without prior treatment, which has a severe negative effect on the environment [30].
Separation technologies with the application of membranes are used successfully and can be considered an integral part of the continuous processing of agro-industrial, food, pharmaceutical and biotechnological products [31]. RBY treatment processes require productive technologies that are efficient and robust enough to explain the intrinsic variability and sometimes fluctuating availability of by-products throughout the year [32] [33] [34].
Extraction of peptides from RBY requires several steps of chemical or mechanical transformation of the cell wall and proteolytic hydrolysis to ensure the transformation of proteins into peptides [35]. The resulting yeast extract contains several macro and micronutrients that must be separated correctly before applying them as new ingredients. Thus, having a more purified product, with higher protein content and fewer unwanted secondary substances, the hydrolysate requires further processing. The separation and fractionation of yeast proteins can be performed by chromatographic methods that have high selectivity, but very high operating costs [36] [37] [38].
The potential of reuse and transformation of this material has been addressed by several authors in an attempt to reduce environmental impact of beer production and promote the recovery of a nutrient-rich by-product [37] [38] [39].
Electroactivation of biological media has a non-residual process, without the use of chemical reagents and relatively inexpensive, which allows the fractionation and obtaining of high quality products [40].
The aim of the research was to simultaneously obtain two products-protein concentrates from residual brewer's yeast (PCRBY) and hydrolyzed protein from residual brewer's yeast (HPRBY) by the electroactivation method.

Materials and Methods
The fresh residual brewer's yeast was collected at JSC "VITANTA", Chisinau, All tests were performed in triplicate. Results were expressed as mean ± standard deviation (SD) and were statistically evaluated (p < 0.05).

Results and Discussion
Undiluted RBY electroactivation, according to configuration 1, allowed the extraction of protein fractions in PCRBY of approximately 93% -94% in the first 5 -10 min of processing. Towards the end of the process (20 -25 min) there was an increase in the protein content (HPRBY, approximately 10%) (Figure 3).
The protein profile of RBY consists of low molecular weight proteins (caseins, albumin), which are subjected to hydrolysis and form peptide-rich hydrolysates.
The variation of pH values and redox potential (E, mV) at undiluted RBY electroactivation characterizes the multiple redox processes that take place in the cathode cell, including sedimentation at the isoelectric point (pI) (Figure 4).
The processing of undiluted RBY was difficult, conditioned by the formation of several active complexes, which tend to migrate through the membrane, which led to its intense soaking, confirmed by the rapid increase in voltage.
Dilution of RBY (1:3) allowed more intense hydrolysis of protein content and extraction of about 16% protein in HPRBY ( Figure 5). Food and Nutrition Sciences   The faster increase of the pH values and, respectively, the decrease of the redox potential values are an obvious indication of the multiple processes that take place at the electroactivation of diluted RBY, confirming the intensification of the dilution process by increasing the activation surface ( Figure 6). The low values of the redox potential highlight the reduction reactions, which lead to the hydrolysis of proteins and, respectively, the obtaining of hydrolysates.
Diluted (1:5) RBY electroactivation (configuration 3) allowed more intense hydrolysis of proteins, thus obtaining in the liquid phase a mixture of protein hydrolysates almost three times larger for the end of processing in the liquid phase of CC (Figure 7).
The pH values towards the end of the process are intensely alkaline, creating favorable conditions for obtaining protein hydrolysates from RBY. Negative values of redox potential (E, mV) at 15 -20 min of processing (C3), confirm the reduction reactions to diluted RBY electroactivation. This can be explained by increasing the activation area ( Figure 8).
The obtained results demonstrate that the electroactivation of RBY allows obtaining two fractions: protein concentrates and protein hydrolysates. Multiple mechanisms require confirmation by supplementation by electrophoretic research of protein concentrates from residual brewer's yeast (PCRBY) and hydrolyzed protein from residual brewer's yeast (HPRBY).
The intense flow of calcium ions through heterogeneous MK-40 membranes in the anode cell allows the ennobling of protein concentrates and hydrolysates with these ions. The presence of albumin in the protein fractions of RBY allows the formation of protein complexes with calcium, attributing a higher biological value to the obtained products.

Conclusions
RBY electroactivation allowed the separation of fractions and the obtaining of protein concentrates and protein hydrolysates. In the fractionation of peptides and proteins, the effects associated with electroactivation and processing parameters were taken into account, which allowed the recovery of economically viable peptides. The integration of load-based separation techniques as well as the elucidation of the underlying separation mechanisms can improve the efficiency of bioseparations.
Several possibilities involve the recovery of value-added compounds from RBY. The production of fractions with a high content of peptides, but with a low content of polysaccharides and fibers is a great challenge.
Preventive dilution of RBY allowed to increase the activation surface and quantitatively intensified the obtaining of protein hydrolysates. The flow of calcium ions through the heterogeneous membrane ennobles the products obtained by electroactivation with these ions.

R. Tarna et al. Food and Nutrition Sciences
Recovery of different fractions enriched in different high value-added components, such as peptides for different applications, oligosaccharides, minerals and amino acids is possible through multiple fractionation processes. This can increase the economic viability of the RBY processing.