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Diwakar Singh

Document Type

Thesis - University Access Only

Award Date


Degree Name

Master of Science (MS)

Department / School

Dairy Science

First Advisor

Sanjeev K. Anand


Membrane fouling is a major operational problem that leads to reduced membrane performance and premature replacement of membranes. Bacterial biofilms developed on reverse osmosis membranes cause severe flux decline during whey process1ng. Our previous studies provided clear evidence of mixed species biofilms on the membranes obtained from a commercial whey processing plant. Presence of resistant microflora such as Bacillus, Escherichia coli, Klebsiella, Enterococcus, Streptococcus, Staphylococcus, Micrococcus, Aeromonas, Corynebacterium, and Pseudomonas species was established. The present investigation was thus undertaken to evaluate a typical CIP protocol against membrane biofilm isolates in planktonic and embedded states. The CIP protocol tested against the constitutive microflora of membrane biofilms included six treatment steps based on; alkali, surfactant, acid, enzyme, a second surfactant, and a weekly sanitizer application. Single and mixed species biofilms developed under in vitro static and dynamic conditions were treated with individual steps of existing CIP protocol. The results obtained from the chemical treatments against 24h old biofilms confirmed the higher effectiveness of acid treatment against biofilm embedded cells amongst all the CIP chemicals. Bacillus isolates showed the highest resistance in planktonic, as well as, biofilm embedded states. Among all the five isolates of Bacillus, isolate (5.10.3) of 10 mo. old consortium was identified as the most resistant. Studies based on the application of sequential CIP protocol against all the consortia indicated survivors even after the complete cleaning process including the sanitizer treatment step. This study thus helped to conclude that the existing CIP protocol was not effective to completely remove biofilms developed on the membrane surface. The study aimed to remove membrane biofilms by developing improved cleaning strategies, especially the enzyme cleaning step. β-galactosidase enzyme was observed to be the most effective under static and dynamic conditions. In addition, a .Protease and a Lipase were also observed to be effective against biofilms developed using the most resistant Bacillus isolate (5.10.3). Studies related to the CIP modifications using β-galactosidase enzyme (CIP-1) revealed lower efficacy as compared to the existing CIP protocol against single and mixed species biofilms. Similarly, the surfactant replacement trials using Span 85, and Tween 85 also did not result in any improvements in the existing CIP protocol. The second approach (CIP-2) by modifying the cleaning conditions (pH and time) concluded better biofilm removal as compared to the existing CIP protocol. Similarly, combination of protease, lipase, and β-galactosidase enzymes and the modified cleaning conditions used as the third approach (CIP-3) indicated higher log reductions against single and mixed species biofilms as compared to the CIP-1 and CIP-2 treatments.

Library of Congress Subject Headings

Reverse osmosis
Membranes (Technology)


Includes bibliographical references (pages 80-102)



Number of Pages



South Dakota State University


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