Types of Ethanol

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Protocols for understanding different types of ethanol and when to use them.

Note: this protocols page may be outdated.

Denatured ethanol vs non-denatured ethanol

The main difference is that denatured ethanol contains additives that make it unsafe to drink. On the other hand, non-denatured ethanol is ethanol with no additives or denaturants.

Denatured ethanol is a form of ethanol containing a high amount of additives and denaturants that makes it poisonous. Denatured ethanol has bad taste and foul smell that makes it unsafe for drinking. There are two forms of non-denatured ethanol named 95% ethanol and absolute ethanol. These forms are known as “pure” forms of ethanol.

Histological/reagent ethanol vs molecular grade

Histological/reagent ethanol is denatured ethanol. Molecular grade ethanol is non-denatured. Source

Use 95% Pure (190 proof) non-denatured Ethanol without Benzene for Preserving tissues for genomic DNA

This article has the following information on ethanol for tissue preservation:

  • If possible, the use of non-denaturated ethanol is favoured, but traces of denaturating agents (e.g. isopropanol, acetone, methyl ethyl ketone, also gasoline) usually do not significantly alter the performance.
  • According to plenty of tests, the optimal concentration is about 95–99% (King and Porter 2004, who also compared with isopropanol).
  • With 70% ethanol, a high degree of degradation has been observed, even after one year.
  • Interestingly, absolute ethanol may not be the optimal medium as it is most expensive and can contain traces of benzene that seriously affect DNA preservation (Ito 1992).
    • Side note: KEL tried to find Ito 1992, but the orginal print is not available online.
  • The ethanol-tissue ratio should exceed 5:1, although some authors recommend much higher ratios (e.g. Martin 1977).
  • When ordering, look for “pure ethanol” and check the SDS to make sure it does not contain additives
  • 95.6% ethanol is an azeotrope, which means the vapor state has the same ethanol-to-water ratio as the liquid state. This means as it evaporates the remaining liquid will still be at 95.6%
  • When adding wet tissues to 95% ethanol, it can decrease the concentration of ethanol. It can be helpful to change the ethanol after the first few months.
  • More information on other ways to preserve tissues are found in this article

ORDERING INFORMATION:Ethyl alcohol, Pure 190 proof, meets USP testing specifications, non-denatured ethanol 493538-4L

Ethanol, 190 proof, 95%, Reagent ACS, Spectrophotometric Grade, Thermo Scientific Chemicals AC615110040

USP vs. ACS ethanol: ACS reagent grade ethyl alcohol meets or exceeds the specifications the American Chemical Society’s Reagent Handbook. USP Grade are chemicals manufactured under current Good Manufacturing Practices (cGMP) and which meet the requirements of the US Pharmacopeia. ACS and USP are generally equivalent.

Use 70% Denatured Ethanol for Cleaning

Information from this article explains the use of ethanol for cleaning. The main points are summarized here:

  • Although it may seem counter-intuitive, 70% ethanol seems to be the most effective concentration, as 90% ethanol coagulates the protein, and the protein layer that is created effectively renders the cell dormant. At a 70% concentration, however, the ethanol takes longer to evaporate and this longer contact time means that it is able to penetrate cells more effectively. The water in a 70% grade is also important in denaturing proteins. And 70% ethanol also tends to be cheaper and is less flammable.

There are a couple of things to watch out for when diluting ethanol:

  • First, diluted ethanol is no longer an azeotrope. At 70%, ethanol will evaporate faster than water, so evaporative loss will result in a gradual decrease in ethanol concentration in the remaining solution. Don’t be surprised when an unsealed beaker of 70% ethanol ‘stops working’. So don’t make more than is necessary at a time, and make sure to label the date when it was made (which you should do for everything you make anyway).
  • Second, ethanol/water mixtures don’t have additive volumes. That means that if you want to make 1 liter of 70% ethanol from absolute ethanol, you can’t do it by mixing 700 ml of ethanol and 300 ml of water. The correct method is to measure 700 ml of ethanol and then bring it to a volume of 1 liter with water.
    • Why? When you mix water and ethanol, the ethanol molecules occupy the space between the water molecules. A helpful way to visualize this is to think about sand and rocks – if you were to mix 1 liter of each, the final volume would be less than 2 liters because the sand would fall between the rocks.

Use 100% (200 Proof, absolute ethanol, molecular grade) non-denatured Ethanol for Molecular Biology work

  • Something about the process of making ethanol makes it hard to get from 95% to 100%. Sometimes this contains additives to get it up to 100%, but they are not denaturing agents.
  • Some procedures that are sensitive to the presence of water require absolute ethanol.
  • A common method to produce ethanol with a higher concentration than 95% is to use additives that disrupt the azeotrope composition and allow further distillation. For this reason, absolute ethanol sometimes contains trace amounts of these additives (such as benzene). source

ORDERING INFORMATION Ethyl alcohol, Pure 200 proof, for molecular biology E7023-4L