Gas Chromatography in DUI Cases – Theory and Operation

By Patrick T. Barone

*From the upcoming 2010 update of Barone’s book, Defending Drinking Drivers

The process of gas chromatography involves the use of an instrument called a gas chromatograph (GC) to separate and analyze compounds that can be vaporized without decomposing the compound. Gas chromatography is particularly well suited to the separation of volatile organic compounds.

Human blood is a mixture of various substances and left alone, it is very difficult to analyze. However, some of the components of blood are volatile organic compounds, and the point of GC is to separate and analyze the volatile organic compounds that may be within the blood sample.

Thus, with the proper extractions procedures or for volatiles such as alcohols by use of a method called “head space,” various foreign components of the blood such as drugs, drug metabolites, and alcohols in blood can be measured and identified. In a drunk driving case we are primarily interested in the volatile ethyl (beverage) alcohol, but there are other potential volatiles of interest, such as acetone, which might be of interest where the driver was experiencing a diabetic episode.

Like all chromatographic methods, with gas chromatography there is a “mobile phase,” in this case a gas, which is used to carry the mixture over a “stationary phase.”  The gas is more fully called “headspace gas.”  With drunk driving cases, the stationary phase is typically a tube or capillary column.  The components in the mixture containing a driver’s blood leave this column in the order of their volatility, with the most volatile (first to vaporize) leaving the column first.

In the forensic lab testing blood for DUI cases, the gas chromatographic system might include the following:

  • The blood sample
  • The headspace vial
  • The internal standard
  • The carrier gas
  • The capillary column
  • The “oven”
  • The flame ionization detector (FID)
  • The computer
  • The printer

The capillary column is contained within an oven. The headspace gas is injected into the column and is measured as it come out of or “elutes” from the column.

Before an unknown volatile can be measured, it is important for the lab analyst to prepare a standard mix which usually includes several different volatiles including isopropyl alcohol, ethyl alcohol, methanol, acetone, acetaldehyde and toluene. This standard mixture allows the laboratory to determine the specific retention times of the various volatiles of interest.

Once the known standards mixture is tested with a specific column and the retention times recorded, the lab analyst can then use the gas chromatograph to qualitatively test unknown compounds.

A calibration curve is also produced in the laboratory.  This involves passing known quantities of alcohol through the column. The specific laboratory’s protocol will dictate how many different levels of alcohol are measured, but they will usually span from well below the legal limit to well above.  This calibration curve “tests” the column to be sure that it is capable of measuring specific known quantities.  Subtle changes in carrier gas flow; the flow of gases to the FID detector (if used) and subtle changes in the column are few of the reasons that the GC must be recalibrated very frequently.

Now that the column has been calibrated, the lab analyst is ready to begin the blood test.  The analyst starts by removing a very small amount of the driver’s blood from the blood draw vial and placing it, along with a very small amount of an internal standard, into a separate testing vial. This testing vial is called a “headspace” vial.  Internal standards are alcohols that would not be expected to occur in human blood except in minute quantities.  More importantly, the boiling point of these standards is different from the boiling point of ethanol.  This difference is important because it will result in a GC peak for the internal standard that is clearly distinguishable from the peak for ethanol. The internal standards that are typically used include n-propyl and t-butyl alcohol.

This headspace vial is then shaken to mix the chemicals and heated to produce the headspace gas. An injector system is used to introduce the sample to be tested into the GC column.  In the case of headspace method, a small amount of air (gas) above the liquid in the headspace vial which has become saturated with volatile components from the liquid sample is taken and injected into column via a micro syringe.  On its way into the headspace vial, the syringe passes through a rubber gasket.  With the headspace method no blood is directly being sampled because headspace testing involves an analysis of only the air above the blood sample.

Once the blood sample has been heated to produce the headspace gas, this gas is swept into and through the column by the carrier gas stream.  This phase is known as the mobile phase. A high pressure gas cylinder serves as the source of the carrier gas. There are several carrier gases that can be used, including helium, nitrogen and hydrogen.

Today, columns used in forensics are generally capillary columns up to 30 meters in length and are made of glass. The diameters of these columns are generally in the range of 0.25 mm. Modern capillary columns typically do not have “packings” as they once did but instead have a coating deposited onto the internal wall of the column. The capillary column consists of a solid support phase and a bonded liquid phase. In capillary columns, the solid support phase is the column itself.

The capillary column separates the sample into its component parts. The oven helps control the speed and amount of separation. The detector detects the presence of and can measure the amount of the volatiles as they exit out of the column. A common detector used in GC systems is the flame ionization detector (FID). However there are many other detectors that are used for special detection requirements.

The FID is located at the end of the column. Because the volatiles involved are flammable, they can be burned in the flame ionization detector.  Thus, as the chemicals exit or “elute” from the end of the column the FID incinerates them, and this combustion produces an electronic charge in the form of ions.

These ions are then measured by the detector and subsequently converted by the instrument’s computer into a graph which usually contains two peaks. One of these peaks represents the internal standard and the other the ethyl alcohol.  The retention time of the peak for the ethyl alcohol must match the expected retention time in order to qualitatively confirm its identity. The expected size and retention time of the peak for the internal standard will be known because of prior testing and because a precise amount of it was placed by the analyst into the headspace vial.  The area beneath this peak, called the “area under the curve” is compared with the peak for the ethyl alcohol.  This ratio is compared with the calibration curve and converted into the driver’s blood alcohol level.

Swimming Pool Metaphor

Perhaps some of the qualitative aspects of GC blood testing can be more fully understood metaphorically.  Think of an Olympic swimming competition where at the beginning of the competition all the swimmers are anonymous. Your goal in this fictitious competition is to figure out the identity of the swimmers. As usual, at the beginning of the race the swimmers all begin at the starting line and as the race begins the swimmers jump into the pool and quickly begin to separate as they race toward the finish line.

While you watch the competition unfold the same event is repeated over and over again and you see the same swimmers compete against each-other several times.  As you watch you begin to notice that the each individual swimmer seems to finish each race at the same time.  If you were keeping notes of each individual swimmer’s time, pretty soon you’d be able to identify the swimmer based on the amount of time it took him or her to finish the race.

Similarly, in the world of GC, the volatile organic compounds finish the “race” at different rates depending on their various chemical and physical properties and their interaction with a specific column.  The volatiles are qualitatively identified based on the amount of time it takes to finish the “race” to the GC’s detector. In gas chromatography this is what happens when the lab analyst runs the standards mixture and records the time the various volatiles elute from the column. This is the qualitative part of gas chromatography, and answers the question “what am I.”

To carry our metaphor a step further, the swimming pool would be similar to the column in the GC.  The solid phase is a bit like the water in the swimming pool in that the water creates a resistance against which the swimmers must race through as they continue toward the finish line.  This resistance helps to separate out the fast swimmers from the slow swimmers, and this separation makes it easier to determine the swimmer’s identity.


1 Response to “Gas Chromatography in DUI Cases – Theory and Operation”

  1. 1 DUI Blood Testing by Gas Chromatography | Michigan DUI Help Trackback on November 12, 2009 at 6:35 pm

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