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What, Where, and How of Adding Antibiotics

Antibiotics Help Control Microbial Contamination in Ethanol Production

This article is based on a presentation by Dr. Dennis Bayrock, Department of Microbiology and Food Science, University of Saskatchewan, Canada. Bayrock (306-211-5779) spoke in June 2004 at the Fuel Ethanol Workshop & Trade Show in Madison, WI.

In his presentation, "Control of Microbial Contamination in Fuel Alcohol Production," Bayrock covered the stresses on yeasts, the types of contaminants in ethanol plants, and the What, where, and how of antibiotic addition.

Yeast Stress

Yeast cells face stress from nonmicrobial (sugar content, sulfite, temperature, sodium and CIP chemicals), microbial (acectic acid, ethanol, lactic acid, pH, and mycotoxins), and nutritional (Lack of sterols, nitrogen, oxygen, UFA, and minerals / vitamins) factors.

Most contaminants at ethanol plants fall within two categories:

Bacteria: Latobacillus sp. and Acetobacter / Ginconobacter sp.
Wild yeasts: Brettanomyces spp; Saccharomyces pasterianus / Bayanus; and Saccharomyces cervisia.

The good news is that controlling bacterial contamination is possible. The differences between culture yeast and bacteria are great enough that they can be exploited during production.

The bad news, however, is that there is no effective control strategy yet for wild yeast. There is promising research in the nutritional differences between yeasts and osmotic tolerances, though.

The best current control practice is proper sanitation and a clean-in-place strategy. Identifying yeasts can be difficult because they are all eukaryotes (organisms with nucleated cells). Morphology, carbohydrate profile, and DNA analysis are the primary ways to identify yeasts.

Antibiotic Dosing — What to Add

It's important to know what you are targeting; hard data is necessary. Selective and differential plant counts are the best methods to get that data. Know the type and number of organisms, and do not rely on high performance liquid chromatography (HPLC) data alone.

Be familiar with antibiotics, including stability, cost, effectiveness, and availability. Know the layout of the ethanol plant, its process flows, and where substances could be added.

Antibiotic considerations. There are several facts to consider before using antibiotics in an ethanol plant.

Cost: Consider the cost per dose rather than the purchase cost.
Stability in system: For example, after 0.8 hours, Penicillin will have 50% remaining at a pH level of 4 and temperature of 40 degrees C. After six hours, 80% of Virginiamycin will remain at a pH level of 4 and a temperature of 63 degrees C.
Residuals in DDGS: Most plants rely on distillation and drying of distillers dried grains (DDGS) as the last steps to destroy any remaining active antibiotics.
Effectiveness: Low doses should be used, the antibiotic should be active against organisms in the plant, and no resistance / adaptation should occur in the use of antibiotics.

Antibiotics: The antibiotics most used at ethanol plants fall into two main categories: Penicillin-based and Virginiamycin-based.

Penicillin Advantages

It is cheap to produce and formulate in various antibiotic mixtures.
Penicillin VK is more pH stable.
It is temperature and pH unstable, which is good for DDGS production.
There are sources everywhere.
It is soluble in water and easy to add.

Penicillin Disadvantages

It is only active against cells that are multiplying, so cells that are already present are not affected.
It only affects Gram+ve bacteria.
It is temperature and pH unstable, so it does not last long in sacch tanks or fermenters.
Microbial resistance is easy to acquire.
Worker's could be allergic to the antibiotic.

Virginiamycin Advantages

It is temperature and pH stable, so it is good for process addition at various placed in the plant.
It is nontoxic to workers.
It is effective against both multiplying and non-multiplying bacteria.
Microbial resistance is hard to acquire.

Virginiamycin Disadvantages

It only affects Gram+ve bacteria.
It is soluble in ethanol, but only slightly soluble in water.

See the chart for which antibiotics are the most effective during different stages of bacteria growth.

Many different details need to be considered when constructing antibiotic dosage and effectiveness charts, including; location of addition; antibiotic stability at location; antibiotic mode of action; type of antibiotic; contact time, and bacterial "load".