br Conflict of interest br Introduction Milk and

Conflict of interest
Introduction Milk and dairy products are nutritious foods that provide lactose, calcium, phosphorous, magnesium and proteins, among other nutrients, which have been reported to have a fundamental role in bone development and healthy diets. An adequate consumption of dairy products throughout life can protect the human body against debilitating bone diseases, such as osteoporosis (Euromilk, 2016, The Dairy Council, 2016). The content of lactose in cow milk is close to 5%, but it cannot be absorbed as such. It must be hydrolyzed into its saccharides components by the action of intestinal β-Galactosidase, an enzyme that transforms lactose into glucose and galactose, two sugars which are readily absorbed into the bloodstream (Mlichová and Rosenberg, 2006). Unfortunately, many Ketoprofen are unable to enjoy the benefits of milk due to intolerance or even allergy to lactose. Approximately 70% of the human population has a reduced ability to digest lactose after childhood, and the frequency of lactose intolerance varies considerably between different ethnic groups (Genetics Home Reference, 2016, The Dairy Council, 2016). This inability to digest lactose is caused by a deficiency of the enzyme β-Galactosidase in the digestive tract (Horner et al., 2011). Apart from the use of the enzyme to offer alternatives to lactose intolerant consumers, lactose hydrolysis can form galacto-oligosaccharides (GOSs), which favor intestinal bacterial microflora (Mlichová and Rosenberg, 2006). It can also be used to produce high sweetness syrups (Horner et al., 2011). Moreover, monosaccharides formed by lactose hydrolysis are fermented more efficiently, reducing the time to obtain fermented products, such as cottage cheese and yogurt. Incorporating principles of sustainable development to improve the environmental profile of a biotechnological process such as enzyme production requires the comprehensive assessment of the production scheme. The biotechnological process is often considered as an example of an environmental friendly system, offering benefits over more conventional chemical-based alternatives. The methodology of Life Cycle Assessment (LCA) is the most widely used environmental tool to assess the environmental impacts of production processes for its entire life cycle (ISO 14040, 2006). Specifically, in the topic of enzyme production, a few reports deal with the environmental impact of industrial production; however, the inventory data reported is limited and potentially subjected to confidentiality issues (Nielsen et al. 2007a, 2007b; Kim et al., 2009). In particular, no LCA reports are available in the literature regarding the production of the β-Galactosidase enzyme. In the framework of designing environmentally sustainable diets (Coelho et al., 2016), this study reports the LCA outcomes of the β-Galactosidase production for lactose hydrolysis from a cradle-to-gate perspective. Considering that this enzyme is essential in the formulation of lactose free products, this paper aims to develop a detailed life cycle assessment of the enzyme production, that can be included in the analysis of food for lactose intolerant consumers. Among different alternatives of production process of the β-Galactosidase enzyme, the use of recombinant Saccharomyces cerevisiae yeast expressing the lacA gene of Aspergillus niger was considered due to its better production potential in comparison with wild strains (Domingues et al., 2004).
Materials and methods
Interpretation of results and discussion
Conclusions The LCA of β-Galactosidase production provided valuable information to evaluate the environmental performance together with data quality from each system. Cleaning-in-Place operations required to prepare the equipment for the next production batch has been identified as the environmental hotspot mainly due to the significant requirement of chemicals, involving contributing ratios to the global environmental stresses ranging from 75% to 92%. Therefore, future improvement actions should be focused on finding alternative chemicals. In fact, a sensitivity analysis has been proposed and reductions of up to 70% could be expected in some impact categories, considering H2SO4 instead of HNO3. Moreover, the chemicals used in the downstream processing (SS2) led to significant impacts (mainly Tris HCl and NaOH required in ion exchange column). Regarding SS1, this subsystem reported negligible contributions to the global environmental profile except for WD, due to the large requirement of water for the preparation of the culture medium. The range of environmental results reported in this study must be carefully compared to other ones available in the literature since depending on the enzyme, the production system requires complex and intensive downstream processing activities coming into larger impacts.