Discussion

The stability of casein micelles in milk systems is highly influenced by the concentration of ethanol and acid (Horne, Reference Horne, McSweeney and O'Mahony2016; Ye and Harte, Reference Ye and Harte2013). In our study, LO made with milks at pH values of 6.7 and 6.0 were turbid (Tables 1 and 2; online Supplementary Fig. S1), probably associated with insufficient levels of ethanol-acidity to completely induce destabilisation of caseins (online Supplementary Fig. S2). Lower levels of ethanol in LO made at pH 6.0 were associated with a reduced perception of alcohol when analysed by sensory analysis (Table 2).

Differences of colour found in LO samples could be attributed due to a decrease in turbidity values as level of milk acidification increased (Table 1). Since a white background was placed for colour measurements, samples with reduced turbidity probably reflected more light from background, leading to higher whiteness (L* values) when compared to those treatments with high turbidity. The green (−a* values) and yellow notes (+b* values) found in all LO samples can be attributed to the presence of riboflavin in the serum phase of milk (Fox et al., Reference Fox, Uniacke-Lowe, McSweeney and O'Mahony2015). The yellowish appearance of LO can also be associated with the addition of saffron, a spice usually used to increase yellowness in food products (Raina et al., Reference Raina, Agarwal, Bhatia and Gaur1996).

The sensory whiteness of LO had an opposite effect when contrasted with instrumental colour analysis (L* values). This could be explained due to differences on how these parameters were measured, since sensory analysis was performed using a transparent cup and instrumental colour measurements were made using a white background. Increased perception of creaminess in LO treatments made with reduced acidity could be associated with higher levels of total protein (Table 2). We believe that certain volatile compounds found in milkfat, such as lactones, give a particular milk flavour to LO samples (Fox et al., Reference Fox, Uniacke-Lowe, McSweeney and O'Mahony2015), although its perception could have been reduced due to a masking effect caused by extensive acid addition in those treatments made with low pH values (Table 2). Nevertheless, flavour perceptions found in LO are also characteristics from other ingredients, such as vanilla and cloves. Hence, the appearance, mouth feeling, flavour and aroma of LO are highly influenced by the addition of milk and non-milk ingredients.

In contrast with caseins, the presence of whey proteins in all treatments of LO (online Supplementary Fig. S2) can be used as authenticity indicator to verify that the final product was originally made from milk, since these components are not affected by the stability of milk ethanol/pH (Garcia Botaro et al., Reference Garcia Botaro, Real de Lima, Aquino, Rocha Fernandes, García and Veiga dos Santos2007; Horne, Reference Horne, McSweeney and O'Mahony2016). Other soluble milk components that may also be found in LO are lactose and Ca. Thus, the presence/absence of proteins, sugars and minerals from milk can be used as indicators to determine authenticity of LO.

In conclusion, a minimum of milk acidification and addition of ethanol is required for destabilisation of milk proteins that leads to LO with a typical transparent appearance. In addition, LO contains combined milk and non-milk components that can be used as markers to certify its authenticity. This is the first study that reports the chemical, physical and sensory properties of LO and can be used as a basis for further research to determine its quality in commercial products.