**1. Introduction**

Agro-industrial waste and by-products streams occur in each step of the food supply chain, specifically during processing. These streams, however, still contain compounds of importance to develop further exploitation schemes, considering also the transition from a linear to circular bioeconomy. Likewise, cheese whey (CW) corresponds to an unavoidable by-product stream of the dairy industry, receiving critical attention because of the high environmental burden, but also owing to the several components with beneficial nutritional and functional properties [1,2]. The compositional analysis of the onset material usually outlines the deployment of subsequent valorization routes within a biorefinery concept to generate high added-value products along with zero waste. For instance, up to date, the vast majority of studies related to the utilization of CW through bioconversion processes implement the application of microbial entities able to consume lactose [3–6]. As a result, the range of end-applications, particularly sustainable food production, is restricted. Alternatively, whey lactose fraction could be hydrolyzed to the respective monosaccharides and

further studied in fermentation processes. Apart from the conventional chemical methods for lactose hydrolysis, previous studies have also undertaken enzymatic hydrolysis [7,8].

Lactose hydrolysis is accomplished via the action of galactosidases, which are ubiquitous enzymes with complex structures. Galactosidases confer several advantages in food industry, including the manufacture of lactose-free dairy products or galacto-oligosaccharides synthesis through transglycosylation reactions [8,9]. Bacterial, yeast and fungal strains correspond to microbial sources of β-galactosidase (β-gal; EC 3.2.1.23 commonly known as lactase), attracting significant interest owing to the ability to secrete the enzymes extracellularly along with featuring properties such as high catalytic activity and reaction rate [10]. On top of that, environmentally benign enzyme production using crude renewable resources as low-cost media has been demonstrated by several species [11]. Notably, several *Aspergillus* species constitute key producers for sustainable and cost-effective enzymes production, also classified as "generally recognized as safe" (GRAS) by the Food and Drug Administration [12]. Currently, evidence for β-galactosidase production exists in the closely related strains of *Aspergillus lacticoferratus* and *Aspergillus awamori* [13,14]. In particular, *A. awamori* produces various hydrolytic enzymes such as glucoamylase, protease, phytase, β-glucosidase, β-xylosidase and cellulases useful for agro-industrial by-product-stream valorization [15–18].

The development of effective and feasible consolidated biorefining should include raw materials with consistent composition, yearlong supply and engage the holistic exploitation of each valuable compound for further novel applications. Extensive studies have been performed to utilize CW derived lactose for the fermentative production of several microbial metabolites [19]. Equally, the protein fraction prevailed in studies targeting novel food formulations [19]. However, the ideal concept would encompass the valorization of both protein and lactose fractions within the same biorefinery approach. Likewise, targeted intermediate products (e.g., biodegradable polymers) within a biorefinery process could be used as onset materials to elaborate "de novo" diversified novel formulations. Bacterial cellulose (BC) is a natural extracellular polysaccharide demonstrating prominent food and biomedical applications, also characterized as GRAS dietary fiber by the FDA in 1992 [20]. Numerous research studies have suggested the use of BC in food applications, including as a flavor additive, fat replacer, stabilizer, rheology modifier and meat analog [20]. Few recent studies also indicated the use of BC as an edible carrier for cell cultures, enzymes, antimicrobial compounds or even biocolorants [21–23]. Despite the simple downstream processing steps, industrial BC production is hindered owing to the high cost of conventional synthetic media. Therefore, agro-industrial by-products and food waste streams have been previously assessed as fermentation supplements for cost-effective BC production [24–27].

Our ultimate target is to develop a holistic approach to exploit cheese whey fractions to generate value-added products, with potential food formulations. Likewise, this initial study describes a two-stage bioprocess to produce crude β-galactosidase and proteases using *A. awamori*, followed by enzymatic hydrolysis of whey lactose, to formulate a nutrient rich feedstock. BC was selected as a case study of an intermediate value-added product. The optimization of crude enzymes production and enzymatic hydrolysis was undertaken via the assessment of several crucial parameters that affect enzyme secretion (e.g., pH value, temperature, enzyme loading). The performance of enzymatic hydrolysis was also assessed, and the obtained hydrolysate was subsequently evaluated as a crude nutrient supplement to generate BC.
