Princeton University Environmental Health and Safety
The Waste-Paper
The Hazardous Waste Disposal Monthly Update
Volume 6 Issue 2 February
2003
Ethidium Bromide
Ethidium bromide is commonly used in Princeton University laboratories.
The purpose of this article is to outline the hazards of ethidium bromide,
precautions and protective equipment that may reduce the risk, and disposal
options.
Hazards
Ethidium bromide is strongly mutagenic and is irritating to the eyes, skin,
mucous membranes and upper respiratory tract. The health effects
of ethidium bromide exposure have not been thoroughly investigated.
It is suspected to be carcinogenic and teratogenic because of its mutagenicity,
although there is no direct evidence of either effect.
The toxic effects of ethidium bromide may be experienced if swallowed,
inhaled or absorbed through the skin.
Safe Handling
Preparation of stock solutions and any operations capable of generating
ethidium bromide dust or aerosols should be conducted in a fume hood to
prevent inhalation.
Gloves should be worn at all times. Latex gloves offer little
protection from ethidium bromide -- nitrile gloves are much more effective
When working with high concentrations or for a prolonged period of
time, double gloving can further reduce the risk of exposure, especially
if the outer glove is replaced whenever significantly contaminated.
Spills or Personal Exposure
As with any chemical, if spilled on skin or eyes, rinse for 15 minutes
using a safety shower or eyewash. If inhaled or swallowed, seek medical
attention immediately at McCosh Health Center or Princeton Medical Center.
For spills, use a spill pillow or absorbent to soak up aqueous ethidium
bromide. Carefully clean up solid ethidium bromide to avoid creating
dusts. Place in a sealable container and dispose in a medical waste
box.
Waste Disposal
Electrophoresis Gels
Trace amounts of ethidium bromide in gels should not pose a hazard.
EHS recommends the following:
-
Less than 0.1% ethidium bromide: place in laboratory trash
-
More than or equal to 0.1%: place in biohazard box for incineration.
(pink or red)
Ethidium Bromide Solutions
Solutions containing ethidium bromide should be filtered, or deactivated,
neutralized and poured down the drain with copious amounts of water.
Deactivation may be confirmed using UV light to detect fluorescence.
There are three recognized methods for deactivation:
Armour Method
This is the simplest method, but is somewhat controversial. One study
found traces of mutagenic reaction mixtures using this method.
1. Combine equal amounts of ethidium bromide solution and household
bleach.
2. Stir constantly for four hours or let sit for 2-3 days.
3. Adjust pH to 4-9 with sodium hydroxide.
4. Pour down drain with copious amounts of water.
Lunn and Sansone Method
For each 100 ml of ethidium bromide solution:
1. Add 5% hypophosphorus acid.
2. Add 12 ml of 0.5 M sodium nitrate.
3. Stir briefly and let stand for 20 hours.
4. Adjust pH to 4-9 using sodium hydroxide.
5. Pour down drain with copious amounts of water.
Quillardet and Hoffnung Method
This method uses 0.5 M potassium permanganate and 2.5 M hydrochloric acid.
Since chlorine gas may be released in significant concentration, EHS does
not recommend using this method.
Charcoal Filtration
Filtering the aqueous ethidium bromide waste solutions, free of other contaminants,
through a bed of activated charcoal is a relatively simple and effective
method for removal of ethidium bromide. Schleicher and Schuell (603-352-3810
or http://www.s-and-s.com) supply filter funnel kits that use packaged
charcoal disks that are graduated for easily tracking the amount of aqueous
solution calculated for a fixed quantities of ethidium bromide residue.
After filtering the ethidium bromide solution, the filtrate is poured down
the drain and the filter is disposed in a biohazardous waste box.
As an alternative, the Green Bag® Kit allows rapid concentration
of ethidium bromide from large volumes of solutions into a small "tea"
bag containing activated carbon which is then disposed as biohazardous
wastes. One kit has the capacity to remove 500 mg of ethidium bromide from
solutions (10mgEtBr/bag). Green Bags are available through BIO101
(www.bio101.com) or through VWR (www.vwr.com - this is a non-catalog item
- use #BIO101 22350-200).
Gloves, Equipment and Debris
Gloves, test tubes, paper towels, etc., that are grossly contaminated with
ethidium bromide should be placed in medical waste for incineration.
Consider deactivating in bleach before disposal if the items are significantly
contaminated.
For further information on this and other waste disposal topics, see the EHS
web page.
Drain Disposal of Chemical
Waste
The Environmental Health and Safety Office administers a Hazardous Waste
Disposal contract that provides a safe and legal means for the proper disposal
of chemical waste through EPA permitted waste disposal companies.
The argument is often made that it is acceptable to dispose of almost
anything down the drain because it will be diluted by several orders of
magnitude before it reaches the sewer treatment facility. Although
this principle of dilution is valid, the waste may first come in contact
with undiluted waste from a nearby lab before it leaves the building.
This may result in a hazardous reaction or may simply expedite the deterioration
of building pipes.
The University does not charge the department nor individual users for
any of the costs of routine chemical waste disposal. The decision not to
charge back these costs was made, in part, to avoid providing an incentive
for improper disposal via laboratory hoods or the sanitary sewer.
The Stony Brook Regional Sewerage Authority (SBRSA) has stringent rules
concerning drain disposal of:
· "known toxic compounds"
· commercial solvents
· flammable liquids, solids, or gases
· corrosives
· metals
· cyanides
The SBRSA samples sanitary sewer lines immediately downstream from Princeton
University to monitor compliance with the requirements.
These restrictions, in addition to New Jersey Pollutant Discharge Elimination
Rules, effectively preclude drain disposal of most wastes. There
are a few wastes for which either drain disposal via the sanitary sewer
or disposal via laboratory trash is safe and permissible. These are
included in the list of Non-Hazardous Chemical Wastes on the EHS website
(http://www.princeton.edu/~ehs/nonhazlist.html) For
any other wastes, do not dispose via the drain without express permission
from EHS. These include aqueous solutions that are biodegradable,
water soluble, free of disagreeable odors, and non-flammable, and water-soluble
inorganic chemicals composed of low toxicity anions and cations.
NEVER dispose of chemical waste via a storm sewer.
Take a close look at the procedures for handling chemical wastes in
your laboratory or work area and avail yourself of the existing chemical
waste disposal system. Seek ways to minimize wastes as much as possible.
If you have specific questions about waste disposal contact Robin Izzo
or Steve Elwood.
Is the Waste-Paper a waste of paper?
If you would like to receive the Waste-Paper electronically, contact Marcia
Leach at 8-5296 or mailto:marcians@princeton.edu.
Hazardous Waste Contacts
at EHS:
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