BACTERIAL CONTAMINATION IN ETHANOL FERMENTATION

KENNETH M. B ISCHOFF, S IQING L IU, T IMOTHY D. LEATHERS, RONALD E. WORTHINGTON, JOSEPH O. R ICH.NAT IONAL CENTER FOR AGRICULTURAL UT IL IZAT ION RESEARCH, U.S. DEPARTMENT OF AGRICULTURE. PUBL ISHED 2008 WILEY PERIODICALS, INC DOI 10 .1002/BIT.22244

PRESENTED BY: MOHAMMAD ALI AL-AMEENNOVEMBER 29 2012BIOPROCESSING AND BIOTECHNOLOGYDR. SONIA M. TIQUIA-ARASHIRO

THE BIG THREE..What is ethanol? • Ethanol is a colorless volatile flammable liquid. Ethanol is also

called ethyl alcohol, pure alcohol, grain alcohol, or drinking alcohol.

How is it produced? • Fermentation is when certain species of yeast (e.g.,

Saccharomyces cerevisiae) break sugar in reduced-oxygen conditions to produce ethanol and carbon dioxide.

Why it’s produced? • It’s the main component of many important commercial

products such as Alcoholic beverages, food, drugs, and also used as an energetic fuel component.

THE PROBLEMBacterial contamination is a critical problem, particularly in fuel ethanol fermentations that are not performed under sterile, pure-culture conditions.

• Bacteria would grow on the substrate available for the yeast yielding competition with yeast and lower production of ethanol.

• Infections could occur and the fermenting facility may result in a “stuck” fermentation which require a “costly” complete shutdown of the facility.

• The emergence of antibiotic resistant bacteria may limit the effectiveness of antibiotics to treat bacterial contamination, and therefore, there need to be a tool that test the development of new antibacterial agents.

• This tool should also monitor the growth of contaminants in the medium.

EMERGING SOLUTIONS?

What criteria are followed in this model?• Cost-efficiency• Effective• Simple

SHAKE FLASK FERMENTATION• Bioreactors are good for simulating the industrial fermentation

process, but their high cost in terms of equipment, media consumption, and labor limit the practical number of combinations of these variables.

• The author decided to monitor the contamination using a shake-flask fermentation model challenged with bacterial strains isolated as contaminants of fuel ethanol facilities.

• Strains of Lactobacillus harvested from ethanol fermentation facilities will be used as the challenging contaminant.

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PUTTING THE MODEL INTO EXAMINATION

The goal is to model bacteria growth and reaction to antibiotics. However, there are variables that needs to be examined including:

• The feedstock• YP/glucose and corn mash

• Bacterial Strain• Six lactic acid bacteria

• Bacterial load• Amount of bacteria added (testing between 104 CFU/ml to 108 CFU/ml).

• Antibacterial treatment.• Treating with Virginiamycin

HPLC ANALYSISHigh performance liquid chromatography was used to determine the concentrations ethanol, glucose, lactic acid, and acetic acid.

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RESULTS AND DISCUSSION

THE FEEDSTOCK

Use of glucose in YP media may represent the state of a chronic bacterial infection, but doesn’t effectively simulate the state of acute infections.

BACTERIAL STRAIN

L. Fermentum 0315-1, L.fermentum 0315-25 and L.brevis 84 produced the most deleterious effect on the yeast-catalyzed fermentation

BACTERIAL LOAD

TREATMENT WITH ANTIBIOTIC

L.Fermentum 0315-1 didn’t get affected by Virginiamycin

CONCLUSIONBacterial contamination in ethanol fermentation is a serious problem that needs to be monitored. In this article, shake-flask fermentation is used. Several variables were studied on this model.

• Not enough variables. There are more variables that can be looked at like Nitrogen and Potassium in the media which might cause fermentation to stop if not present in a good amount

• Is this model better than Bioreactor monitoring? • They used HPLC which can be substituted by the bioreactor itself

monitoring the growth and concentration itself.• How does this model work on sterile, pure culture conditions. Is it more

accurate than the bioreactor?• Industry can afford a bioreactor.

REFERENCES• Partitioning of Potassium during Commercial-Scale

Red Wine Fermentations and Model Wine ExtractionsAm. J. Enol. Vitic. March 1, 2009 60:43-49

• Influence of Medium Buffering Capacity on Inhibition of Saccharomyces cerevisiae Growth by Acetic and Lactic Acids

Appl. Environ. Microbiol. April 1, 2002 68:1616-162• Effect of yeast inoculation rate on the metabolism of

contaminating lactobacilli during fermentation of corn mash.• J Ind Microbiol Biotechnol. 2004 Dec;31(12):581-4. Epub

2004 Dec 14

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