Efficient removal of spores from skim milk using cold microfiltration: Spore size and surface property considerations

Bacterial spores present in milk can cause quality and shelf-life issues for dairy products. The objectives of this study were to evaluate the effectiveness of microfiltration (MF) in removing Bacillus licheniformis and Geobacillus sp. spores from skim milk using membranes with pore sizes of 1.4 and 1.2 µm, and to investigate the role of spore surface properties in MF removal. Cell sizes were determined by scanning electron microscopy, surface charge by zeta potential analysis, and surface hydrophobicity by contact angle measurements. Commercially pasteurized skim milk was inoculated with a spore suspension at about 10⁶ cfu/mL, and then processed by MF using ceramic membranes at 6°C, a cross-flow velocity of 4.1 m/s, and transmembrane pressure of 69 to 74 kPa. Total aerobic plate and spore counts in the milk were determined before and after MF. All processing runs and surface and product analyses were conducted in triplicate, and data were analyzed statistically. For the same strain, spores were shorter and wider than vegetative cells, averaging 1.37 to 1.59 µm in length and 0.64 to 0.81 µm in width. Reduction of B. licheniformis spores significantly increased with a reduction in MF pore size, from 2.17 log for 1.4-µm pore size, to 4.57 log for 1.2-µm pore size. Both pore sizes resulted in almost complete removal of Geobacillus sp. spores. All spores and the ceramic membrane had a negative surface charge at milk pH, indicating an electrostatic repulsion between them. Bacillus licheniformis spores were hydrophilic, whereas Geobacillus sp. spores were hydrophobic. Consequently, Geobacillus sp. spores had a tendency to cluster in skim milk, preventing their passage even through the 1.4-µm MF membrane, whereas some B. licheniformis spores could still pass through a 1.2-µm membrane. This study demonstrates that efficient removal of spores from skim milk by cold MF may require a smaller membrane pore size than required for removal of vegetative cells of the same species, and that cell surface properties may interfere with the outcome of filtration as would be anticipated based on size alone.