# Chemical-Sensory Traits of Fresh Cheese Made by Enzymatic Coagulation of Donkey Milk
## 1 Introduction  
The use of equid milk for human nutrition and wellness has an ancient history. The use of mare milk for producing koumiss, a lactic-alcoholic beverage, belongs to the food tradition of several ethnic groups from Central Asia [1]. Ancient Romans and Greeks were already aware of the cosmetic and therapeutic properties of donkey milk, as evidenced by the legendary milk bath of [PERSON] and [PERSON], and [PERSON] indications as a remedy for several diseases [2, 3]. After a long and slow decline, donkey rearing is now undergoing a revival, due to an interest in innovative and nutraceutical foods. Donkey milk is not in competition with cow's milk, and represents an added value functional food with high market potential. Besides, European policy encourages the diversification of agricultural production and the valorisation of local genetic resources. Donkeys are able to use marginal areas unsuitable for specialised dairy cows, and they are better adapted to warm climates.  
For these reasons, rearing donkeys for milk is gaining increasing attention in the Mediterranean area, in particular Italy [4], where milk is sold as a raw product directly at farms. Other forms of marketing at niche level also exist, such as selling online or through pharmacies as pasteurised or, rarely, as UHT-treated or freeze-dried product [5]. The interest of the food researchers towards donkey milk is also increasing; several protocols for manufacturing innovative fermented mills and functional beverages at laboratory level are already available [6; 7; 8]. Unfortunately, studies on cheesemaking are very rare, since rennet coagulation is commonly considered as unfeasible. Apart from economic considerations connected to the high cost and low availability of the raw matter, the main technical obstacles seem to be the low casein concentration (0.64 to 1.03 g/100 g milk) and different micelle structure with respect to ruminants [9]. Recent studies have reported that donkey casein micelles are much larger than bovine micelles (298.5 \(\pm\) 18.9 nm at pH 6.8 versus 186.5 \(\pm\) 1.2 nm), have a lower absolute zeta potential (\(-15.4\pm 0.5\) mV), poor colloidal stability and different abundance of the casein fractions [10; 11]. This latter aspect has long been debated, until the work of [PERSON] et al. [12] definitively clarified the matter using electrophoresis and immunostaining of the protein bands with polyclonal antibodies against the four 'classic' caseins. The authors ascertained the presence of \(\kappa\)-casein, and demonstrated that \(\beta\)-casein was the most abundant fraction, followed by \(\alpha\)s1 and \(\alpha\)s2.  
Despite the difficulty in milk coagulation, some successful efforts in making fresh donkey cheese have been recently reported. In particular, [PERSON] [13] and [PERSON] et al. [14] developed two different cheesemaking protocols, based on the use of camel chymosin or calf rennet, respectively. Even though the results obtained were encouraging, the two studies had some conflicting points, and the products were only poorly investigated. The aim of the present study was to provide a more in-depth insight on the chemical and sensory traits of a cheese made from donkey milk by using microbial rennet.