Evaluating the Effectiveness of Individual Cleaning Steps of a CIP Protocol in Membrane Biofilm Removal Under Dynamic Conditions

This study evaluated the effectiveness of individual clean-in-place (CIP) steps in removing biofilms from reverse osmosis (RO) membranes under dynamic flow conditions using the Centers for Disease Control (CDC) biofilm reactor. Biofilms were developed in the laboratory under continuous flow, using mixed-species bacterial isolates obtained from 10-month-old RO membrane biofilms from a commercial facility. Individual CIP chemicals, representative of those used in commercial protocols, were tested against 24 h-old biofilms. Additionally, a complete six-step sequential CIP process was conducted under dynamic conditions, consisting of treatments with alkali, surfactant, acid, enzyme, a secondary surfactant, and sanitizer. All experiments were performed in quadruplicate, and data were subjected to statistical analysis. Among individual treatments, the acid step was the most effective, significantly outperforming the other CIP cleaning steps by reducing bacterial counts from 5.62 to 4.10 log units, a 96.98% reduction. The full six-step CIP protocol reduced counts to 2.24 log units, indicating the persistence of resistant cells. The presence of viable cells post-treatment highlights the limited efficacy of the tested CIP chemicals in fully eradicating mature biofilms. Additionally, skipping any step in the membrane cleaning can significantly compromise the efficiency and performance during production. These findings suggest that biofilms grown in vitro under dynamic conditions using the CDC reactor exhibit a more robust assessment of the CIP treatments in accomplishing the biofilm control. This study highlights the need for optimized, scientifically validated CIP protocols targeting biofilms to improve cleaning efficacy and food safety.