Role of mono- and oligosaccharides from FOS as stabilizing agents during freeze-drying and storage of Lactobacillus delbrueckii subsp. bulgaricus

Abstract

The aim of this work was to assess the role of mono- and oligosaccharides present in fructo-oligosaccharides (FOS) mixtures as protective agents during freeze-drying and storage of Lactobacillus delbrueckii subsp. bulgaricus CIDCA 333. Different FOS mixtures were enzymatically obtained from sucrose and further purified by removing the monosaccharides produced as secondary products. Their glass transition temperatures (Tg) were determined at 11, 22 and 33% relative humidity (RH). Bacterial cultures were freeze-dried in the presence of 20% w/v solutions of the studied FOS. Their protective effect during freeze-drying was assessed by bacterial plate counting, and by determining the lag time from growth kinetics and the uptake of propidium iodide (PI). Plate counting during bacterial storage at 4 °C, and 11, 22 and 33% RH for 80 days completed this rational analysis of the protective effect of FOS. Purification of FOS led to an increase of Tg in all the conditions assayed. Microorganisms freeze-dried in the presence of non-purified FOS were those with the shortest lag times. Bacteria freeze-dried with pure or commercial FOS (92% of total FOS) showed larger lag times (8.9–12.6 h). The cultivability of microorganisms freeze-dried with non-purified FOS and with sucrose was not significantly different from that of bacteria before freeze-drying (8.74 ± 0.14 log CFU/mL). Pure or commercial FOS were less efficient in protecting bacteria during freeze-drying. All the protectants prevented membrane damage. The cultivability of bacteria freeze-dried with FOS decayed < 1 logarithmic unit after 80 days of storage at 11% RH. When storing at 22 and 33% RH, pure and commercial FOS were those that best protected bacteria, and FOS containing monosaccharides were less efficient. The effect of FOS on bacterial protection is the result of a balance between monosaccharides, sucrose and larger FOS in the mixtures: the smallest sugars are more efficient in protecting lipid membranes, and the larger ones favor the formation of vitreous states.