Freeze-drying is commonly used to preserve probiotics, but it could cause cell damage and loss of viability. The cryoprotectants play an important role in the conservation of viability during freeze-drying. In this study, we investigated the survival rates of Lactobacillus reuteri CICC6226 in the presence of cryoprotectants such as sucrose, trehalose, and reconstituted skim milk (RSM). In addition, we determined the activities of hexokinase (HK), pyruvate kinase (PK), lactate dehydrogenase (LDH), and ATPases immediately following the freeze-drying. The results showed that the differences in HK and PK activities with and without the cryoprotectants during freeze-drying were not significant, but cell viability and activities of LDH and ATPase were significantly different (P<0.01) prior to and after freeze-drying. Meanwhile, the results showed that the maintenance of the membrane integrity and fluidity was improved in the presence of the 10% trehalose or 10% RSM than other treatments during freeze-drying. These results have provided direct biochemical and metabolic evidence of injured cell during freeze-drying. Freeze-drying damaged membrane structure and function of cell and inactivated enzymes (LDH and ATPases). The results imply that LDH and ATPases are key markers and could be used to evaluate the effect of cryoprotectants on viability and metabolic activities of L. reuteri CICC6226 during freeze-drying.
Cheese is a typical handcrafted fermented food in Kazak minority from the Uighur Autonomy Region in China and Central Asia. Among the microbial community that is responsible for Kazak cheese fermentation, yeasts play important role in flavor formation during ripening. To develop ripening cultures, we isolated 123 yeasts from 25 cheese products in Kazak, and identified 87 isolates by the D1/D2 domain of the large subunit rRNA gene sequence. Pichia kudriavzevii was the dominant yeast in Kazak cheese, followed by Kluyveromyces marxianus and Kluyveromyces lactis. Of these, the ability to exhibit enzyme of dominant isolates and contribution to the typical flavor of cheeses was assessed. Enzyme producing yeast strains were inoculated in Hazak cheese‐like medium and volatile compounds were identified by head space solid phase micro extraction coupled to gas chromatography and mass spectroscopy. Pichia kudriavzevii N‐X displayed the strongest extracellular proteolytic and activity on skim milk agar and produced a range of aroma compounds (ethanol, ethyl acetate, 3‐methylbutanol, and acetic acid) for Kazak cheese flavor, could be explored as ripening cultures in commercial production of Kazak cheeses.
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