Through­out this series of posts we have exam­ined the Thorton-Nakumura pro­to­col that is used through­out the United States for the pros­e­cu­tion of ille­gal pos­ses­sion mar­i­juana. A fair exam­i­na­tion of the ques­tion reveals that there is no valid­ity to the notion that the 3 test reg­i­men pro­duces a valid con­clu­sion that the unknown exam­ined in fact con­tains THC.

Here are those series of posts:

  1. What is the goal and the pur­pose of test­ing of unknowns gen­er­ally? How do we best design a test for marijuana?
  2. How is most mar­i­juana test­ing con­ducted in the United States?
  3. What is micro­scopic mor­pho­log­i­cal exam­i­na­tion? Is it a “good” test?
  4. What is the mod­i­fied Duquenois-Levine test? Is it a “good” test?
  5. What is Thin Layer Chro­matog­ra­phy? Is it a “good” test?
  6. Is the com­bi­na­tion of all three tests cre­ate a “good” test­ing scheme?
  7. Is there a bet­ter way to test for marijuana?

There is a bet­ter way: Mod­ern Instru­men­ta­tion

There is an instru­ment dri­ven way that is very spe­cific and pro­duces ver­i­fi­able data. This instru­ment dri­ven tech­nique has been val­i­dated and if use in the val­i­dated man­ner with a prop­erly trained oper­a­tor uti­liz­ing proper sam­ple col­lec­tion, proper sam­ple selec­tion, proper sam­ple prepa­ra­tion, per­fect instru­men­ta­tion, and legit­i­mate inter­pre­ta­tion of the data will arrive at a valid result. This mod­ern day instru­men­ta­tion is called the Gas Chro­matog­ra­phy with Mass Spec­trom­e­ter (GC-MS).

Gas Chromatography with Mass Spectrometer

Gas Chro­matog­ra­phy with Mass Spectrometer

When the lab­o­ra­tory ana­lyst is pro­fi­ciency tested on unknowns and then graded to see whether or not they can con­duct a proper analy­sis of an unknown, the ana­lyst uses GC-MS. When the lab­o­ra­tory wants to check that the known that they have pur­chased from a third party ven­dor that is used and needed in Thin Layer Chro­matog­ra­phy (TLC) to com­pare against the unknown, the lab­o­ra­tory requires that it be ver­i­fied by GC-MS.

So when the lab­o­ra­tory really wants to know or really needs to know whether or not some­thing con­tains delta 9 THC, it uses the most spe­cific device avail­able that pro­duces ver­i­fi­able data. The ver­i­fi­able data are the print­outs that result from the analy­sis. This is called a Total Ion Cur­rent (TIC) chro­matogram and the result­ing spec­trum that is com­pared against an adju­di­cated known that is pro­duced by the National Insti­tutes of Stan­dards and Tech­nol­ogy (NIST).

Here are other blog posts on the GC-MS process:

  1. Introduction-The dif­fer­ent con­fig­u­ra­tions and the Elec­tron Impact process
  2. What types of mass ana­lyz­ers are there?
  3. What type of detec­tors are there?
  4. What types of analy­sis can be done?
  5. How do you read the output?
  6. How do they come to a qual­i­ta­tive mea­sure using software?
  7. How do they quan­ti­tate the results?
  8. Do you need chro­matog­ra­phy if you are using Mass Spectrometry?
  9. Other top­ics of inter­est about GC-MS

In the analy­sis of  unknowns that are seized, the process of deriva­ti­za­tion can be used to volatilize the sam­ple for intro­duc­tion the the GC-MS. For exam­ple, the ana­lyst can use MTBSTFA (N-methyl-N-(tert-butyldimethylsilyl)-trifluoroacetamide) or BSTFA/TMCS N,O-bis (trimethylsi­lyl) trifluoroacetamide/ Trimethylchlorosi­lane or MSTFA: N-methyl-N-trimethylsilyltrifluoroacetamide to deriv­a­tive the unknown.

A pop­u­lar tech­nique includes:

A sam­ple prepa­ra­tion that includes 500 mg of dry and homog­e­nized herbal cannabis are extracted with 5 ml methanol : chlo­ro­form (9:1 v/v) by the fol­low­ing pro­ce­dure: 10 sec­onds on a vor­tex, 15 min. ultra­sonic bath includ­ing again vor­tex­ing after 5, 10 and 15 min­utes, then cen­trifu­ga­tion. The sam­ple then needs to go through decar­boxy­la­tion. A 200 μl of the above extract are trans­ferred into a deriva­ti­za­tion ves­sel. The sol­vent is evap­o­rated under nitro­gen gas to dry­ness. The sam­ple is decar­boxy­lated for 15 min­utes at 210°C. The residue is dis­solved in 200 μl methanol : chlo­ro­form (9:1 v/v). The prepa­ra­tion of the final solu­tion next involves tak­ing the above decar­boxy­la­tion solu­tion and dilut­ing it with methanol by a fac­tor of 100 (in two steps, each 100 μl + 900 μl) and is then used for the analysis.

Also, THC is very much amenable to Head­space Solid Phase Microex­trac­tion (HS-SPME). It is a non-derivatization-based tech­nique where there is a extrac­tion from the solid dose itself. It is a more direct mea­sure as one is not chem­i­cally chang­ing the sam­ple. Dif­fer­ent types of fil­ters and fibers can be used such as poly­di­methyl­silox­ane 100 μm.

GC based test­ing for THC has been dis­cussed in  the sci­en­tific lit­er­a­ture since 1971. In 1991 the UNDCP study dis­cussed both GC-based meth­ods and HPLC meth­ods to iden­tify THC.

Other for­eign gov­ern­ments require much more test­ing than we do here in the United States. For exam­ple, Canada requires four tests includ­ing two instrument-based analy­sis that pro­duce ver­i­fi­able infor­ma­tion such as a spectrum.

There is a bet­ter way, but it is not used. This cer­tainly can­not be fair and just. It is time that we as cit­i­zens demand proof in the court­room and end the myth of spe­cific iden­ti­fi­ca­tion of mar­i­juana in crim­i­nal courtrooms.

The way to close this series is by an apt obser­va­tion made by Dr. Fred­eric White­hurst, PhD JD who wrote:

Many a defense attor­ney has looked in amaze­ment at the court­room “iden­ti­fi­ca­tion” of Cannabis Sativa L. by a law enforce­ment offi­cer sport­ing a gilded pot metal badge and a high school diploma who has writ­ten a report that would turn a ninth grade Eng­lish teacher’s hair gray and deny that same offi­cer a high school diploma. Obvi­ously the award­ing of a Basic Law Enforce­ment Train­ing (BLET) diploma ele­vates such high school grad­u­ates to the doc­toral level in botany, enabling them to con­duct a visual leaf archi­tec­tural analy­sis and the court accepted alchemy of the Duquenois Levine test to reach a con­clu­sion that the green veg­etable mate­r­ial in that lit­tle baggy can be iden­ti­fied as mar­i­juana to the exclu­sion of all other plant material.

The mod­ern day pros­e­cu­tion for the unlaw­ful pos­ses­sion of mar­i­juana is based upon a three test régime involv­ing micro­scopic mor­pho­log­i­cal exam­i­na­tion, mod­i­fied Duquenois-Levine col­ori­met­ric test­ing, and Thin Layer Chro­matog­ra­phy. Each of these three tests are non-specific for THC which is the phar­ma­co­dy­nam­i­cally active ingre­di­ent which makes mar­i­juana ille­gal. The ques­tion becomes is this three test bat­tery col­lec­tively con­clu­sively spe­cific to arrive at a valid con­clu­sion that the unknown seized and tested is in fact mar­i­juana (con­tains THC) and there is no pos­si­bil­ity of a false positive?

In this series of posts we are going to exam­ine this seemly sim­ple question:

  1. What is the goal and the pur­pose of test­ing of unknowns gen­er­ally? How do we best design a test for marijuana?
  2. How is most mar­i­juana test­ing con­ducted in the United States?
  3. What is micro­scopic mor­pho­log­i­cal exam­i­na­tion? Is it a “good” test?
  4. What is the mod­i­fied Duquenois-Levine test? Is it a “good” test?
  5. What is Thin Layer Chro­matog­ra­phy? Is it a “good” test?
  6. Is the com­bi­na­tion of all three tests cre­ate a “good” test­ing scheme?
  7. Is there a bet­ter way to test for marijuana?

Part 6: Is the com­bi­na­tion of all three tests cre­ate a “good” test­ing scheme?

The gov­ern­ment would choose to argue that the com­bi­na­tion of these three tests results in a valid con­clu­sion that this tested unknown is mar­i­juana (con­tains THC). All  three roads leads, so they say, lead to THC. The more sophis­ti­cated ver­sion of this argu­ment is the Venn dia­gram. Per­haps some­thing like the below:

Venn diagram that the prosecution argues
Venn dia­gram that the pros­e­cu­tion argues

This is sim­ply not proven or sup­ported by the pub­lished empir­i­cal research. Even if it were true there is no evi­dence how wide or big that “D” area is. It could be very wide and large con­tain­ing many com­pounds such as this below exam­ple below where the result of this type of test­ing would be a pos­i­tive for O, T, H, P, M, A, B, X, K, Y and not for B alone:

What could be in there? We don't know.
What could it be? We don’t know.

There is no evi­dence that these dis­tinct tests ever intercept.

Maybe these three tests have nothing in common
Maybe these three tests have noth­ing in common

While in ear­lier posts, we have exam­ined the pro­pri­ety and suit­abil­ity of these three sep­a­rate and dis­tinct tests in the iden­ti­fi­ca­tion of THC, we can­not prop­erly judge them in iso­la­tion. We must remem­ber and in fair­ness this is a 3 test process and technique.

There are no mean­ing­ful or robust stud­ies pub­lished that truly val­i­dates this three test pro­ce­dure as result­ing in a spe­cific qual­i­ta­tive mea­sure for THC.

What com­pounds the issue of the pos­si­bil­ity of error is that as the ana­lyst is not using the same sam­ple through­out each of these 3 tests. It is in fact, three dif­fer­ent sam­ples of the orig­i­nal unknown. The ana­lyst pre­sumes that the unknown is homoge­nous. Fur­ther assumed is that the sam­pling and the sam­ple selec­tion of the unknown con­ducted by the ana­lyst result in iden­ti­cal homo­ge­neous sam­ples. These are not jus­ti­fied sci­en­tific assump­tions (An assump­tion is not drawn from evi­dence; it is a hypoth­e­sis {my assump­tion can be tested by look­ing at the dic­tio­nary}. A pre­sump­tion implies a basis in evi­dence {the legal pre­sump­tion of innocence})

Each of these tests are wholly destruc­tive in nature by their very process.

  • Just because the sought after fea­tures of the micro­scopic mor­pho­log­i­cal exam­i­na­tion were present in the first sam­ple doesn’t nec­es­sar­ily mean that that this sam­ple will con­tain the sought after fea­tures in the mod­i­fied Duquenois-Levine and the TLC exam­i­na­tion because they are not exam­ined for in this sample.
  • Just because the sought after fea­tures of the mod­i­fied Duquenois-Levine exam­i­na­tion were present in the sec­ond sam­ple doesn’t nec­es­sar­ily mean that the fea­tures sought after in the micro­scopic mor­pho­log­i­cal exam­i­na­tion are there (because it was not exam­ined) and the sought after fea­tured of the TLC exam­i­na­tion are there (because it was not exam­ined) in this sec­ond sample.
  • Just because the sought after fea­tures of the TLC exam­i­na­tion were present in the third sam­ple doesn’t nec­es­sar­ily mean that the fea­tures sought after in the micro­scopic mor­pho­log­i­cal exam­i­na­tion are there (because it was not exam­ined) and the sought after fea­tured of the mod­i­fied Duquenois-Levine exam­i­na­tion are there (because it was not exam­ined) in this third sample.

All pros­e­cu­tions for the unlaw­ful pos­ses­sion of mar­i­juana requires as an essen­tial ele­ment of the crime for the gov­ern­ment to advance proof that the unknown sub­mit­ted for test­ing is in fact mar­i­juana (con­tains THC). Can the gov­ern­ment actu­ally do that based upon its typ­i­cal test­ing method?

In this series of posts we are going to exam­ine this seemly sim­ple question:

  1. What is the goal and the pur­pose of test­ing of unknowns gen­er­ally? How do we best design a test for marijuana?
  2. How is most mar­i­juana test­ing con­ducted in the United States?
  3. What is micro­scopic mor­pho­log­i­cal exam­i­na­tion? Is it a “good” test?
  4. What is the mod­i­fied Duquenois-Levine test? Is it a “good” test?
  5. What is Thin Layer Chro­matog­ra­phy? Is it a “good” test?
  6. Is the com­bi­na­tion of all three tests cre­ate a “good” test­ing scheme?
  7. Is there a bet­ter way to test for marijuana?

Part 5: What is Thin Layer Chro­matog­ra­phy? Is it a “good” test?

Thin Layer Chro­matog­ra­phy test­ing

What is it?

Thin Layer Chro­matog­ra­phy (TLC) is a chro­mato­graphic tech­nique. It is a com­bi­na­tion of a col­ori­met­ric test and is mea­sured in terms of separation.

Mechan­i­cally how is it preformed?

TLC depends on the sta­tion­ary phase, often a glass plate coated with sil­ica (it must be prop­erly  des­ic­cated or dried) and the mobile phase which is com­prised of a sol­vent mix­ture made of toluene and diethy­lamine typ­i­cally. When placed on a prop­erly dried plate and using a prop­erly mixed and pre­pared sol­vent, the sol­vent (the mobile phase) will be wicked up by the sil­ica with cap­il­lary action and travel up the plate. A sam­ple from the unknown is selected. It is mashed up in some mechan­i­cal process and in some processes call for it to be dis­solved. It is placed in the des­ig­nated spot. A test is com­pared by per­form­ing the reac­tion of the plate of a known sam­ple from an adju­di­cated source. This com­par­i­son is made with the analyst’s eyes. There will be dif­fer­ent spots on the plate at dif­fer­ent posi­tions from the ori­gin with var­i­ous inten­si­ties of color. The height and the color change are visu­al­ized. The spots typ­i­cally need to be visu­al­ized with a chem­i­cal spray such as Fast Blue B Salt (50 mg in 20 ml of NaOH (0.1 N)) or par­tic­u­lar lighting.

How is the typ­i­cal crime lab­o­ra­tory ana­lyst trained to con­duct this?

Once again, the ana­lyst is not trained in the fun­da­men­tals of how or why this process works. If you handed them a pen and paper and ask them to dia­gram and explain with specifics the chro­mato­graphic process, they would likely be baf­fled. Also for­eign to them would be the specifics as to why the sol­vent used to elute up the plate has to be in a spe­cific ratio and not another. In essence, it is another sub­jec­tive test as it is based upon per­cep­tion of color by the ana­lyst and the per­cep­tion of this height devel­op­ing on the plate ver­sus the adju­di­cated known.

The ana­lysts are not aca­d­e­m­i­cally trained in the the­ory of this tech­nique of chro­matog­ra­phy, and are not taught about cross-reactivity and false pos­i­tives or other sources of errors.

Is this a ver­i­fi­able test?

Much like the mod­i­fied Duquenois-Levine test, TLC is poten­tially ver­i­fi­able. Dig­i­tal cam­eras exist. Pho­tographs can be taken. Heck, even video can be taken to show how it is con­ducted on a par­tic­u­lar sam­ple from the unknown and this com­par­i­son to the adju­di­cated known. How­ever, the mod­ern prac­tice is to not take a sin­gle pho­to­graph, to not take advan­tage of video tech­nol­ogy and pro­duce noth­ing ver­i­fi­able in court that the test was even con­ducted or that the analyst’s per­cep­tion of the change did hap­pen and was cor­rectly interpreted.

Again, there are no crime lab­o­ra­to­ries that I am aware of that use the ACE-V (Analy­sis, Com­par­i­son, Eval­u­a­tion, and Ver­i­fi­ca­tion) tech­nique that one would find in fin­ger­print iden­ti­fi­ca­tion with a dou­ble check in real time by a fel­low bench ana­lyst. In essence, it is checked one time, by one per­son with no dou­ble check by another, and noth­ing pro­duced that proves that the analy­sis was done or that the fea­tures that are reported as present were in fact objec­tively present.

Is there empir­i­cal valid­ity stud­ies that prove that this is a spe­cific and con­fir­ma­tory test?

Once again, this type of test­ing has not been proven to be a val­i­dated method to test specif­i­cally for THC. There are well-known and dis­cov­ered false pos­i­tives which includes cof­fee, basil and even tobacco products.

Once again, the same issues that are dis­cussed prove true with this test­ing. This TLC test is pre­formed on a totally dif­fer­ent sam­ple from the unknown that is not sub­jected to the micro­scopic mor­pho­log­i­cal exam­i­na­tion or the mod­i­fied Duquenois-Levine col­ori­met­ric test.

In this series of posts we are going to exam­ine this seemly sim­ple question:

  1. What is the goal and the pur­pose of test­ing of unknowns gen­er­ally? How do we best design a test for marijuana?
  2. How is most mar­i­juana test­ing con­ducted in the United States?
  3. What is micro­scopic mor­pho­log­i­cal exam­i­na­tion? Is it a “good” test?
  4. What is the mod­i­fied Duquenois-Levine test? Is it a “good” test?
  5. What is Thin Layer Chro­matog­ra­phy? Is it a “good” test?
  6. Is the com­bi­na­tion of all three tests cre­ate a “good” test­ing scheme?
  7. Is there a bet­ter way to test for marijuana?

Part 4: What is the mod­i­fied Duquenois-Levine test? Is it a “good” test?

Mod­i­fied Duquenois-Levine test­ing

What is it?

It is tech­ni­cally referred to as a col­ori­met­ric test. In short, it is a color test. A totally dif­fer­ent selec­tion of the unknown is sam­pled and sub­jected to this test. A reagent is added to the unknown. The reagent is made up of a com­bi­na­tion of vanillin, acetalde­hyde, and ethanol in a spe­cific ratio of these com­po­nent materials.

Mechan­i­cally how is it preformed?

This totally sep­a­rate sam­ple from the unknown is placed into typ­i­cally a test tube. A cer­tain amount of the Duquenois reagent is added (typ­i­cally about 10 drops). The tube is closed. The tube is agi­tated (shaken) for an unspec­i­fied period. The tube is reopened. Con­cen­trated hydrochlo­ric acid is then added (usu­ally about 20 drops). The tube is closed. It is agi­tated (shaken) again. Any color change is then noted. The tube is reopened. Chlo­ro­form is added. The tube is closed. It is again agi­tated (shaken) or vor­texed (mixed). The ana­lyst is look­ing for a color change (thought to be vio­let or pur­ple) and a sep­a­ra­tion into two layers.

This is the end result of the modified Duquenois-Levine test. This picture is after the HCl and the chloroform are added. There is a deep purple color at the top and then a pink color at the bottom.

This is the end result of the mod­i­fied Duquenois-Levine test. This pic­ture is after the HCl and the chlo­ro­form are added. There is a deep pur­ple color at the top and then a pink color at the bottom.

How is the typ­i­cal crime lab­o­ra­tory ana­lyst trained to con­duct this test?

They are trained in the process and the steps in the mechan­i­cal process. No crime lab­o­ra­tory ana­lyst is ever instructed by a doc­toral level ana­lyt­i­cal chemist as to why this process results in any sort of color change or the way the phys­i­cal sep­a­ra­tion occurs in the col­ors with the addi­tion of chlo­ro­form or why it these changes hap­pen at all.

In essence, it is a sub­jec­tive test as it is based upon per­cep­tion of color by the ana­lyst and the per­cep­tion of this devel­op­ing of a sep­a­rate layer once chlo­ro­form is added.

It is a sec­ond sam­ple that is tested. Prior to the test­ing by mod­i­fied Duquenois-Levine tech­nique, there is no micro­scopic mor­pho­log­i­cal exam­i­na­tion con­ducted on this sam­ple. There is no TLC analy­sis on this sample.

It is tested one time and then dis­carded. No other test­ing is per­formed on this sample.

The ana­lysts are not aca­d­e­m­i­cally trained in the the­ory of the reagent use, and are not taught about cross-reactivity and false pos­i­tives or other sources of errors.

Is this a ver­i­fi­able test?

Poten­tially, yes. Dig­i­tal cam­eras exist. Pho­tographs can be taken just like the one above. Heck, even video can be taken to show how it is con­ducted on a par­tic­u­lar sam­ple from the unknown. How­ever, the mod­ern prac­tice is to not take pho­tos. They video­tape noth­ing.  The lab­o­ra­tory pro­duces noth­ing ver­i­fi­able in court that the test was even con­ducted or that the analyst’s per­cep­tion of the change in color did hap­pen and was cor­rect or that there was this sep­a­ra­tion once the chlo­ro­form is added.

Again, there are no crime lab­o­ra­to­ries that I am aware of that use the ACE-V (Analy­sis, Com­par­i­son, Eval­u­a­tion, and Ver­i­fi­ca­tion) tech­nique that one would find in fin­ger­print iden­ti­fi­ca­tion with a dou­ble check in real time by a fel­low bench ana­lyst. In essence, it is checked one time, by one per­son with no dou­ble check by another, and noth­ing pro­duced that proves that the analy­sis was done or that the fea­tures that are reported as present were in fact objec­tively present.

Is there empir­i­cal valid­ity stud­ies that prove that this is a spe­cific and con­fir­ma­tory test?

Nope. In fact the empir­i­cal stud­ies clearly show the oppo­site. It is not spe­cific for THC. The reac­tion is not unique to THC. In fact, the stud­ies clearly show that it is not even mean­ing­fully selec­tive for THC. Chi­nese moth­er­wort if tested, by this method will turn vio­let. But that is not all. There are a great many plants that yield sim­i­lar color results when Duquenois-Levine test­ing is applied. Yet ana­lysts are taught that is color change is diag­nos­tic of THC. Naka­mura him­self pub­lished and acknowl­edges that M. J. de Faubert Maun­der listed 25 species of plants which exhib­ited vio­let to pur­ple col­ors in the Duqenois test, and were extractable in chlo­ro­form, which is the dis­tin­guish­ing fea­tures of the Duquenois Levine test. (502) Naka­mura tested 23 of those species noted by M. J. de Faubert Maun­der. Using the Duquenois-Leine test, he found that the vio­let to pur­ple color when test­ing the leaves of the fol­low­ing (502):

  • cof­fee
  • a species of gum copal called Caplafer conjugata
  • gum Kawri
  • wood sage
  • Thuja occi­den­talis
  • San­darac

He pub­lished that he tested and found vio­let to pur­ple color reac­tions using the Duquenois-Levine test in other plants as well, and not just the leafy por­tions of these plants. (502). That list includes:

  • cala­mus
  • cul­ver root
  • gin­ger
  • gum animi
  • gum copal
  • gum myrrh
  • henna
  • let­tuce opium
  • san­dal wood
  • tolu
  • wood betony
  • liquorice
  • nut­meg
  • poi­son flag

The United Nations study on mar­i­juana con­cludes that Duquenois-Levine test on sub­stances com­pletely unre­lated to mar­i­juana will give false pos­i­tives too. This includes Arthemisia Gran­cunulus, Euca­lyp­tus Glab­u­lus Labill and sev­eral other herbs that are very com­mon such as rose­mary, thyme, and sage.

None of these items above con­tain THC. Yet, they fea­ture these color changes and some even have the sep­a­ra­tion when chlo­ro­form is added that the ana­lysts are trained to look for when apply­ing these reagents.

While we have exam­ined the pro­pri­ety and suit­abil­ity of this mod­i­fied Duquenois-Levine test­ing in the iden­ti­fi­ca­tion of THC, we can­not judge this Duquenois-Levine test in iso­la­tion. We must remem­ber and in fair­ness this is a 3 test process and tech­nique. What com­pounds the issue of the pos­si­bil­ity of error is that as the ana­lyst is not using the same sam­ple through­out each of these 3 tests. It is in fact, three dif­fer­ent sam­ples are taken of the unknown and each test gets its own sam­ple with no sam­ple receiv­ing the ben­e­fit of all three tests. The ana­lyst pre­sumes that the sam­pling and the sam­ple selec­tion of the unknown is homo­ge­neous mean­ing that each sam­ple will be exactly iden­ti­cal. This is not a jus­ti­fied sci­en­tific assump­tion (An assump­tion is not drawn from evi­dence; it is a hypoth­e­sis {my assump­tion can be tested by look­ing at the dic­tio­nary}. A pre­sump­tion implies a basis in evi­dence {the legal pre­sump­tion of inno­cence}) Just because the sought after fea­tures of the micro­scopic mor­pho­log­i­cal exam­i­na­tion were present in the first sam­ple, that doesn’t nec­es­sar­ily mean that it will be present in the next sam­ple that the ana­lyst now sub­jects to mod­i­fied Duquenois-Levine testing.

Naka­mura tested and found that mace and nut­meg, which yielded col­ors sim­i­lar to that obtained with mar­i­juana with the Duquenois-Levine test, could be “cred­i­bly con­fused” with mar­i­juana based on micro­scopic appear­ance. (502)

In this series of posts we are going to exam­ine these seemly sim­ple questions:

  1. What is the goal and the pur­pose of test­ing of unknowns gen­er­ally? How do we best design a test for marijuana?
  2. How is most mar­i­juana test­ing con­ducted in the United States?
  3. What is micro­scopic mor­pho­log­i­cal exam­i­na­tion? Is it a “good” test?
  4. What is the mod­i­fied Duquenois-Levine test? Is it a “good” test?
  5. What is Thin Layer Chro­matog­ra­phy? Is it a “good” test?
  6. Is the com­bi­na­tion of all three tests cre­ate a “good” test­ing scheme?
  7. Is there a bet­ter way to test for marijuana?

Part 3: What is micro­scopic mor­pho­log­i­cal exam­i­na­tion? Is it a “good” test?

Micro­scopic mor­pho­log­i­cal examination

What is it?

The micro­scopic mor­pho­log­i­cal exam­i­na­tion in short is an exer­cise of botan­i­cal iden­ti­fi­ca­tion using a microscope.

Mechan­i­cally how is it preformed?

A very small amount of the dried unknown is selected. This becomes the sam­ple. The sam­ple is placed on a micro­scopic slide. A drop or two of water is then added to the slide. The slides are exam­ined at vary­ing lev­els of mag­ni­fi­ca­tion and under dif­fer­ent light con­di­tions. What the ana­lyst is look­ing for is two dis­tinct mor­pho­log­i­cal fea­tures. They are look­ing for micro­scopic “hairs” on the unknown. These are cys­tolithic hairs and glan­du­lar hairs. Cys­tolithic hairs are often likened to like lit­tle bear claws in their appearance.

photomicrographs of cystolithic hairs

pho­tomi­cro­graphs of cys­tolithic hairs

The sec­ond type of hair is called a glan­du­lar hair. These are fre­quently remarked as look­ing like mushrooms.

photomicrographs of glandular hairs

pho­tomi­cro­graphs of glan­du­lar hairs

Some tech­niques call for the use of hydrochlo­ric acid after they look for these hairs. A few drops of HCL are added by the ana­lyst. The ana­lyst then looks to see if there is some unspec­i­fied effer­ves­cence under the light of the microscope.

How is the typ­i­cal crime lab­o­ra­tory ana­lyst trained to con­duct this form of testing?

The ques­tion becomes what expe­ri­ence level in botany and tax­on­omy and microscopy does the ana­lyst truly have? Very few under­grad­u­ate pro­grams exist in botany in the United States. Most ana­lysts have on the job train­ing where another per­son who like­wise have no for­mal train­ing in botany or tax­on­omy that instructs them. It also involves the use of a micro­scope. For­mal train­ing in microscopy is required in order to use a micro­scope prop­erly and to prop­erly inter­pret what the human eye sees through var­i­ous pow­ers and light­ing con­di­tions of the micro­scope. At the end of the in-house train­ing, the typ­i­cal ana­lyst can­not typ­i­cally express the fam­ily, the genus and the species that is “mar­i­juana” or at what power and under what light­ing con­di­tions they saw the mor­pho­log­i­cal characteristics.

This sam­ple that is exam­ined under the micro­scope is then dis­carded. All future or addi­tional test­ing is con­ducted on totally dif­fer­ent sam­ples from the unknown.

Is this a ver­i­fi­able test?

It poten­tially is. There is a device that can be linked to the micro­scope to take pic­tures of what the ana­lyst thinks he or she sees. This is called a pho­tomi­cro­graph. In fact the pic­tures above come from just such a micro­scope that is equipped with one. A dig­i­tal cam­era attached to a micro­scope is very com­monly used in sci­ence. They are very mod­er­ately priced. As they are dig­i­tal cam­eras, the cost of pro­duc­tion and print­ing and data stor­age is neg­li­gi­ble. It is fre­quently used in other types of com­par­a­tive exam­i­na­tions such as some higher lev­els of foren­sic firearm or tool­mark iden­ti­fi­ca­tion. I know of no lab­o­ra­tory in the United States that relies upon micro­scopic mor­pho­log­i­cal exam­i­na­tion that uses mod­ern tech­nol­ogy and pro­duces pho­tomi­cro­graphs. In fact, few, if any, crime lab­o­ra­to­ries use the ACE-V (Analy­sis, Com­par­i­son, Eval­u­a­tion, and Ver­i­fi­ca­tion) tech­nique that one would find in fin­ger­print iden­ti­fi­ca­tion using a stereo-microscope and a dou­ble check in real time by a fel­low bench ana­lyst. In essence, the unknown is checked one time, by one per­son with no dou­ble check by another, and noth­ing is pro­duced that proves that the analy­sis was done or that the fea­tures that are reported as present were in fact objec­tively present. There is no proof.

Is there empir­i­cal valid­ity stud­ies that prove that this is a spe­cific and con­fir­ma­tory test?

No. There are no empir­i­cal and robust val­i­da­tion stud­ies that con­clude that this form of micro­scopic mor­pho­log­i­cal exam­i­na­tion even when the two botan­i­cal fea­tures (cys­tolithic and glan­du­lar hairs) are objec­tively present yield a valid opin­ion that the plant exam­ined is def­i­nitely con­tains THC. There are no stud­ies that say the two fea­tures means that there is THC present. In fact, what stud­ies that are out there con­clude that this form of mor­pho­log­i­cal exam­i­na­tion using a micro­scope is per­haps not even selec­tive. In the orig­i­nal stud­ies by Naka­mura, he indi­cated that cys­toliths of var­i­ous types are found in the leaves of a num­ber of dicots. (497). He also indi­cated that the pres­ence of cys­toliths is not diag­nos­tic for a fam­ily, let alone a genus of plants. (497) Naka­mura specif­i­cally noted that cys­toliths are found on a great num­ber of plants includ­ing but not lim­ited to: hops plants (500), oregano (500), lemon thyme (501), sil­ver thyme (501), and rose­mary (501). Naka­mua he specif­i­cally noted 63 “rep­re­sen­ta­tive” species in 13 plant gen­era that con­tain cys­toliths in table 5 of his arti­cle (501) Naku­mura indi­cated that he made NO attempt to pre­pare a com­pre­hen­sive list­ing of species bear­ing cys­tolith hairs sim­i­lar to those found in cannabis “because of the sheer enor­mity of the task to exam­ine 31,874+ dicotyle­dons.” (500). For instance, in one genus found in Table 5 of his arti­cle, the Loasa, he specif­i­cally noted 9 species that had cys­toliths; how­ever, he went on to say that there were actu­ally some 80 species of that genus known to have sim­i­lar hairs. (501). He fully acknowl­edged that his list­ing was not com­pre­hen­sive. So it is accu­rate and very fair to say that the 63 “rep­re­sen­ta­tive” species that have cys­toliths that were noted by Naka­mura in Table 5 of his arti­cle are not an exhaus­tive list. Other stud­ies agree that at least 6 other sub­stances also have hairs that con­tain these two fea­tures (cys­tolithic and glan­du­lar hairs). In terms of the addi­tional step of adding HCL to the sam­ple and exam­in­ing for effer­ves­cence under the light of the micro­scope, it is quite clear that other sub­stances can pro­duce the same effer­ves­cence when a few drops of hydrochlo­ric acid are added to them. For exam­ple net­tles and cat­nip do exactly that.

Some folks main­tain and tes­tify under oath every day in the United States that this unver­i­fied micro­scopic mor­pho­log­i­cal exam­i­na­tion is diag­nos­tic of iden­ti­fi­ca­tion of THC pres­ence in an unknown. There is no sci­en­tific sup­port for this type of testimony.

One thing that every ana­lyst should agree with is that sim­ply because these hairs are present and if they con­duct HCL addi­tion and if there is effer­ves­cence that does not mean that the unknown con­tains THC. This is why they have to do addi­tional test­ing, mean­ing the mod­i­fied Duquenois-Levine and Thin Layer Chro­matog­ra­phy testing.

What is fre­quently not part of any mor­pho­log­i­cal exam­i­na­tion for cannabis is what botanists have noted to be other fea­tures con­sis­tent with cannabis. The sim­pli­fied exam­i­na­tion for the typ­i­cal foren­sic sci­ence iden­ti­fi­ca­tion is pur­pose­fully designed to make this exam­i­na­tion and con­clu­sions from it eas­ier to per­form by non-botanists. As with every­thing in life, the more cri­te­rion attached to qual­ify some­thing the least likely there will be a qual­i­fi­ca­tion. The exam­i­na­tion of cannabis and espe­cially a mor­pho­log­i­cal exam­i­na­tion by untrained botanists should not be made easy. All of the fea­tures that are known to be diag­nos­tic by the world of botany should be used not sim­ply the easy ones. For exam­ple, botanists have noted that that cannabis has ses­sile glands as well as con­tain­ing ser­rated edges of the leaves and com­pound palmate struc­ture mean­ing sev­eral leaflets arise from the same point. The addi­tion of all of the known mor­pho­log­i­cal fea­tures known to true botanists as diag­nos­tic of cannabis would make this exam­i­na­tion more robust and the result more selec­tive than the sim­plis­tic exam­i­na­tion that now per­me­ates the foren­sic sci­ence world.

In 2006, more than 829,000 peo­ple were arrested in this coun­try for marijuana-related offenses alone. Since 1937 with the pas­sage and adop­tion of the Mar­i­huana Tax Act, mar­i­juana has been effec­tively pro­hib­ited in the United States.

Lit­er­ally mil­lions upon mil­lions of peo­ple have been accused of pos­sess­ing mar­i­juana. The ques­tion becomes are these con­vic­tions sci­en­tif­i­cally supported?

In this series of posts we are going to exam­ine this seemly sim­ple question:

  1. What is the goal and the pur­pose of test­ing of unknowns gen­er­ally? How do we best design a test for marijuana?
  2. How is most mar­i­juana test­ing con­ducted in the United States?
  3. What is micro­scopic mor­pho­log­i­cal exam­i­na­tion? Is it a “good” test?
  4. What is the mod­i­fied Duquenois-Levine test? Is it a “good” test?
  5. What is Thin Layer Chro­matog­ra­phy? Is it a “good” test?
  6. Is the com­bi­na­tion of all three tests cre­ate a “good” test­ing scheme?
  7. Is there a bet­ter way to test for marijuana?

Part 2: How is most mar­i­juana test­ing con­ducted in the United States?

So what hap­pens in Amer­ica, here, now and today in the iden­ti­fi­ca­tion of marijuana?

  • Are the tests pre­sump­tive or confirmatory?
  • Are the meth­ods used verifiable?
  • Does it result in a spe­cific or selec­tive identification?

Let’s take a look at how mar­i­juana is tested in the United States today in the super-majority of cases. It all begins with a police offi­cer or another per­son who seizes an unknown sub­stance. It’s green. It’s veg­etable like. It looks like mar­i­juana to them. Per­haps they do some sort of quick and dirty test on the side of the road using a fast col­ori­met­ric test and there is a change in color. This is a true unknown. We don’t know what it is. We can­not con­clude what it is based upon sim­ply look­ing at it with our own two eyes or even by our sense of smell. This is why we have foren­sic sci­en­tists and fur­ther test­ing because those types of obser­va­tions may lead to an improper or incor­rect result.

attempts at identifying marijuana at the gross or macro level can be misleading

Attempts at iden­ti­fy­ing mar­i­juana at the gross or macro level can be misleading

A basic fun­da­men­tal ques­tion becomes: What makes mar­i­juana ille­gal to pos­sess? What makes mar­i­juana ille­gal is that it con­tains the phar­ma­co­dy­nam­i­cally sub­stance known as Delta-9 Tetrahy­dro­cannabi­nol (THC).

Given that THC is what makes mar­i­juana ille­gal, then log­i­cally what would be best is if we were design tests that react exclu­sively to THC and noth­ing else and pro­duces ver­i­fi­able data that iden­ti­fies that the green veg­etable sub­stance in fact specif­i­cally con­tains THC.

Because we can’t tell what it is just by look­ing at it or just by smelling it, then we must sub­ject this unknown to test­ing. Largely in the United States there is a régime or process of how crime lab­o­ra­to­ries con­duct this test­ing. The triad of test­ing that is con­ducted are:

  1. Micro­scopic mor­pho­log­i­cal examination
  2. Mod­i­fied Duquenois-Levine testing
  3. Thin Layer Chromatography

This is gen­er­ally referred to as the Thorton-Nakumura protocol.

In order to answer our ques­tions of speci­ficity, ver­i­fi­ca­tion, and con­fir­ma­tory we need to look at each of these three test­ing tech­niques. In our next blog posts over the next sev­eral weeks each of these tests will be scrutinized.

In many court­rooms all across the United States today, a state ana­lyst opines under oath that a unknown leafy green veg­etable like sub­stance is in fact mar­i­juana. The ques­tion becomes, in the typ­i­cal case, is this a valid con­clu­sion? Is it the whole sci­en­tific truth?

In this series of posts we are going to exam­ine these seemly sim­ple questions:

  1. What is the goal and the pur­pose of test­ing of unknowns gen­er­ally? How do we best design a test for marijuana?
  2. How is most mar­i­juana test­ing con­ducted in the United States?
  3. What is micro­scopic mor­pho­log­i­cal exam­i­na­tion? Is it a “good” test?
  4. What is the mod­i­fied Duquenois-Levine test? Is it a “good” test?
  5. What is Thin Layer Chro­matog­ra­phy? Is it a “good” test?
  6. Is the com­bi­na­tion of all three tests cre­ate a “good” test­ing scheme?
  7. Is there a bet­ter way to test for marijuana?

Part 1: What is the goal and the pur­pose of test­ing of unknowns gen­er­ally? How do we best design a test for marijuana?

Remem­ber that the goal of all ana­lyt­i­cal test­ing is to pro­duce a valid result. A valid result is the epit­ome of good science.

A valid result is made up of three dis­tinct features:

  1. It is a spe­cific measurement,
  2. using a tech­nique or series of tech­niques that are part of a val­i­dated procedure
  3. that is verifiable.

Fea­tures of a spe­cific test

The first step is that the test­ing must be unique and spe­cific qual­i­ta­tive mea­sure­ment. The dif­fer­ence between speci­ficity and selec­tiv­ity is very impor­tant. To be spe­cific in terms of qual­i­ta­tive mea­sure­ment means that there is an iden­ti­fi­ca­tion of a par­tic­u­lar com­pound and exclu­sively that one spe­cific com­pound to the exclu­sion of every other com­pound or sub­stance in the universe. It is unequiv­o­cal iden­ti­fi­ca­tion based upon objec­tive cri­te­rion and val­i­dated meth­ods or tech­niques that are ver­i­fi­able. In short, it is what it is and could be noth­ing else.

On the other hand, if the iden­ti­fi­ca­tion is selec­tive, then it is not spe­cific. The iden­ti­fi­ca­tion that results from selec­tive test­ing or meth­ods means that it could be that sub­stance but it also could be some­thing else. There could be cross-reactivity, co-elution or just poor sen­si­tiv­ity. There could be false pos­i­tives. In essence and fun­da­men­tally, we do not know what it is defin­i­tively, uniquely, and specif­i­cally. We may not know what it is, but we can learn what it is not.

Gen­er­ally, the dif­fer­ence between a spe­cific test­ing régime and that of a selec­tive one is the dif­fer­ence between con­fir­ma­tory data and pre­sump­tive data and the dif­fer­ence between the sci­en­tific weight of the opin­ions expressed based upon them. A pre­sump­tive test is some­thing that is char­ac­ter­ized by a high rate of false pos­i­tives and cross-reactivity. It is not at all spe­cific and may or may not be use­fully selec­tive. A con­fir­ma­tory test is one that results in a near spe­cific mea­sure­ment with a very, very low rate of false pos­i­tives. A con­fir­ma­tory test or even a com­bi­na­tion of con­fir­ma­tory tests can never be truly 100% cor­rect. There is still room for error. Noth­ing is perfect.

Selective versus Specific is kind of like Sesame Street's "one of these things is not like the other things"

Selec­tive ver­sus Spe­cific is kind of like Sesame Street’s “one of these things is not like the other things”

The Inter­na­tional Union of Pure and Applied Chem­istry (IUPAC), which is the world author­ity on chem­i­cal nomen­cla­ture, ter­mi­nol­ogy, stan­dard­ized meth­ods for mea­sure­ment, atomic weights and other crit­i­cally eval­u­ated data and oth­ers have defined the dif­fer­ence between these often con­fused terms as follows:

A spe­cific reac­tion or test is one that occurs only with the sub­stance of inter­est, while a selec­tive reac­tion or test is one that can occur with other sub­stances but exhibits a degree of pref­er­ence for the sub­stance of inter­est.  Few reac­tions are spe­cific, but many “exhibit selectivity”.

Other com­mon def­i­n­i­tions include:

Selec­tiv­ity gives an indi­ca­tion of how strongly the result is affected by other com­po­nents in the sample.

and also

Selec­tiv­ity refers to the extent to which the method can be used to deter­mine par­tic­u­lar ana­lytes in mix­tures or matri­ces with­out inter­fer­ences from other com­po­nents of sim­i­lar behavior.

A selec­tive test may be not a spe­cific test due to cross-reactivity, inter­fer­ence, coelu­tion, or codetermination.

Fea­tures of a val­i­dated tech­nique or series of techniques

We wrote a whole 6 part series on method val­i­da­tion and what it takes to be valid result.

  1. Part 1: Introduction-Is it valid, invalid or non-validated?
  2. Part 2: What is method validation?
  3. Part 3: Can we use some­one else’s val­i­dated method?
  4. Part 4: What trig­gers ver­i­fi­ca­tion, re-validation or out right new val­i­da­tion of a method?
  5. Part 5: What are the essen­tial terms in method validation?
  6. Part 6: What is qual­ity assur­ance and qual­ity control?

Fea­tures of a ver­i­fi­able test

A ver­i­fi­able test is one in which oth­ers who were not present for the test­ing can look at some­thing to basi­cally dou­ble check what test­ing was pre­formed and its result. With­out ver­i­fi­ca­tion, then there can­not be a val­i­dated result.

The end result in the courtroom

If we, as a soci­ety, sin­cerely mean the prin­ci­ples of the pre­sump­tion of inno­cence, proof beyond a rea­son­able doubt, and the bur­den of pro­duc­tion and per­sua­sion, then in the case of iden­ti­fi­ca­tion of an unknown when it is pre­sented in court and if the gov­ern­ment can only prove selec­tiv­ity, then the jury’s result should always be not guilty.

a typical seized unknown

a typ­i­cal seized unknown

In courts all across the United States evi­dence is pre­sented in the pros­e­cu­tion of those charged with pos­ses­sion (either mis­de­meanor or felony) that the green veg­etable sub­stance seized is in fact mar­i­juana. The way that this is typ­i­cally pre­sented in the court­room is as an absolute and as a spe­cific qual­i­ta­tive mea­sure­ment. This series of posts seeks to ask the ques­tion: Is this in fact true?

In this series of posts we are going to exam­ine this seemly sim­ple question:

  1. What is the goal and the pur­pose test­ing of unknowns gen­er­ally? How do we best design a test for marijuana?
  2. How is most mar­i­juana test­ing con­ducted in the United States?
  3. What is micro­scopic mor­pho­log­i­cal exam­i­na­tion? Is it a “good” test?
  4. What is the mod­i­fied Duquenois-Levine test? Is it a “good” test?
  5. What is Thin Layer Chro­matog­ra­phy? Is it a “good” test?
  6. Is the com­bi­na­tion of all three tests cre­ate a “good” test­ing scheme?
  7. Is there a bet­ter way to test for marijuana?
What the heck is this?

What the heck is this?

The great Bard once penned:

All: God save your majesty!

Cade: I thank you, good people—there shall be no money; all shall eat and drink on my score, and I will apparel them all in one liv­ery, that they may agree like broth­ers, and wor­ship me their lord.

Dick: The first thing we do, let’s kill all the lawyers.

Cade: Nay, that I mean to do.

Henry The Sixth, Part 2 Act 4, scene 2, 71–78

Shakespeare’s char­ac­ter Dick the Butcher’s idea of a per­fect soci­ety was one where jus­tice pre­vailed because there were no lawyers. The trai­tor­ous Jack Cade had not so noble a rea­son for want­ing to get rid of all of the lawyers. He wanted to become the auto­crat in a quasi-communistic social rev­o­lu­tion. Cade alleges that all lawyers do is use laws and lan­guage set up by fel­low lawyers to oppress and ruin the life of every day man. There­fore, in his esti­ma­tion, no jus­tice results.

That is an extreme view.

I sug­gest that per­haps jus­tice best results when we have an edu­cated and orga­nized defense bar who is sci­en­tif­i­cally edu­cated in foren­sic sci­ence. As I have blogged before, the foren­sic sci­ence com­mu­nity as cur­rently prac­ticed in the United States today is very flawed. Extremely flawed. Fun­da­men­tally flawed. In some cases, if not the major­ity of cases, it is utterly unsci­en­tific. I agree with the sen­ti­ments that the crim­i­nal defense com­mu­nity shares a large por­tion of the blame. Many lawyers well before our times, let come into evi­dence prac­tices, tech­niques, and “the­o­ries” that had just but the very veneer of sci­ence and were, how­ever, any­thing but sci­en­tific and far from valid. This insti­tu­tional prop­a­ga­tion of error is a large hur­dle for many of us to over­come now where busi­ness as usual or sim­ple rep­e­ti­tion is some­how equated with validity.

So what are we to do?

Set up pro­grams where we edu­cate the defense bar.

I would like to high­light one of these: The Amer­i­can Chem­i­cal Soci­ety Hands-on Foren­sic Chro­matog­ra­phy course.

It is a five-days hands-on class con­ducted at Axion Ana­lyt­i­cal Lab­o­ra­to­ries, Inc. in Chicago, Illi­nois. This hands-on course is taught by three icons of chro­matog­ra­phy (Dr. Harold McNair, PhD, Dr. Lee Polite, PhD and Mr. Lew Fox) and two attor­neys who spe­cial­ize in eval­u­at­ing chro­matog­ra­phy and foren­sic sci­ence related cases (Justin J. McShane and Josh D. Lee).

 

The agenda includes:

Day 1

8:15am Reg­is­tra­tion and Snacks
8:30am Intro­duc­tion to Gas Chromatography
9:30am Inlet Sys­tems for Liq­uid Injections
10:30am Lab 1: GC Famil­iar­iza­tion and Parameters
11:45 Lab Review
12:00pm Lunch
1:00pm Fun­da­men­tals of Sep­a­ra­tion — Resolution
2:30pm GC The­ory
3:30pm Cap­il­lary Columns
4:30pm Lab 2: Col­umn Installation
6:00pm End of Session
7:30pm Group Din­ner

Day 2

8:30am Quan­ti­ta­tive Analysis
9:30am Dis­cus­sion of QC in the Foren­sic World
10:30am Lab 3: GC Quantitation
12:00pm Lunch
1:00pm Lab Review
2:00pm Head­space GC
4:00pm Lab 4: Head­space Demo
5:00pm End of Session
6:30pm Axion Labs spon­sored Group Dinner

Day 3

8:30am GC-MS The­ory
10:30am LC-MS The­ory
12:00pm Lunch
1:00pm Labs 5–7: Wet Lab –Sam­ple Preparation/ Inte­gra­tion / GC-MS Instrumentation
3:00pm FID Detec­tor
4:00pm Dis­cov­ery
5:30pm End of Session
7:05pm Sport­ing event

Day 4

8:30am Ethics
9:30am Lab 8: Walk­ing Down a Case-How to Sort, Iden­tify and Exam­ine Data (Con­fronta­tion Clause)
11:00 am Gen­eral Review of Major Concepts
12:00pm Lunch
1:00pm Defenses That Work
3:00pm Lab 9: Trou­bleshoot­ing GC Problems
4:30pm Lab 10: Beers and Data Round­table (Bring your own data set eval­u­ate with your small group)
8:30pm End

 

Day 5

8:30am Trou­bleshoot­ing Lab Review
9:00am Cross exam­i­na­tion of an Analyst
10:30am Direct Exam­i­na­tion of an Expert
12:00pm Trou­ble Shoot­ing Lab Review
12:30pm Soft stop of the course with gen­eral discussion
2:00pm Hard Stop-End of Course

This class attracts full of attor­neys from all across the United States. In this class atten­dees not only are instructed in the class­room the­ory that under­lies chro­matog­ra­phy (both liq­uid and gas chro­matog­ra­phy) in gen­eral and the spe­cific the­o­ries that allow for head­space analy­sis and how Flame Ion­iza­tion Detec­tor, UV-DAD and Mass Spec­trom­e­try (EI and EC based) works, and get to see the instru­ments, but they also get to do the fol­low­ing with their own two hands and more:

Hands-on Lab 1

Instru­ment Famil­iar­iza­tion where the atten­dees pre­form direct injec­tions into a Gas Chromatograph-Flame Ion­iza­tion Detec­tor (GC-FID) using EtOH sam­ples, and learn to inter­pret GC-FID chro­matograms, import and manip­u­late a method, change the dif­fer­ent vari­ables on the GC-FID and elu­ci­date the results.

Hands-on Lab 2

The atten­dees on their own GC machine remove and install cap­il­lary columns and all of the com­po­nents of the injec­tor (septa, liner, gold seal, etc.), check for effi­cien­cies, res­o­lu­tion, the­o­ret­i­cal plates, and learn about split ver­sus split­less injec­tor settings.

Hands-on Lab 3

The atten­dees on their own GC machine estab­lish a cal­i­bra­tion curve from CRMs for EtOH. The atten­dees on their own GC machine ana­lyze the response and pro­gram a cal­i­bra­tion curve (exter­nal stan­dard) and also use the Inter­nal Stan­dard method to assure qual­ity. The atten­dees on their own GC machine estab­lish meth­ods and report­ing of this cru­cial part of testing.

Hands-on Lab 4

The atten­dees con­tinue to use Head­space Gas Chromatograph-Flame Ion­iza­tion Detec­tor and also use Gas Chro­matog­ra­phy Mass Spec­trom­e­try (GC-MS) sys­tem, and use an High Per­for­mance Liq­uid Chro­matog­ra­phy (HPLC) system.

Hands-on Lab 5

Sam­ple Preparation-sampling ver­sus sam­ple selec­tion is demon­strated. The atten­dees do their own Pipet­ting. The atten­dees do their own con­trol chart­ing. The atten­dees use vol­u­met­ric flasks. The atten­dees learn about pre-analysis error hands-on and how it affects quan­tifi­ca­tion. The atten­dees are intro­duced to issues of metrol­ogy and Uncer­tainty Measurement.

Hands-on Lab 6

Inte­gra­tion. The atten­dees on their own Chem­Sta­tion work­sta­tion learn about inte­gra­tion and how easy it is to manip­u­late the data. The atten­dees manip­u­late their own data. The atten­dees learn what to look for that shows that the data was manip­u­lated, and the atten­dees learn how to manip­u­late data so that it is not dis­cov­er­able with­out the raw com­puter data in the soft­ware files.

Hands-on Lab 7

GC-MS Instru­men­ta­tion. The atten­dees see how easy life is for an ana­lyst. The atten­dees get to see the analy­sis of the raw data on a GC-MS and dis­cover how a true novice can turn into an “expert” with a sim­ple push of a but­ton. The atten­dees see with their own two eyes the “hid­den” data that the Gov­ern­ment and its lab­o­ra­tory doesn’t want any­one to see that will reveal the truth that the sup­posed “gold stan­dard” that pro­vides for the alleged “unequiv­o­cal iden­ti­fi­ca­tion” of test­ing of unknowns that is GC-MS is not perfect.

Hands-on Lab 8

How to Obtain Discovery/Walking Down a Case/Defenses that Win-Advanced Issue Spot­ting: In this lab, the best prac­tices in how to obtain dis­cov­ery with a spe­cial empha­sis on how to strate­gi­cally and prac­ti­cally build a record so that the atten­dees can get the raw data in its un-manipulated raw com­puter form and also in its print form. The atten­dees go through an actual case that was lit­i­gated that fea­tured experts on both sides, and issue spot all of the prob­lems with the dis­cov­ery and the data as it was presented.

Hands-on Lab 9

Trou­bleshoot­ing. The atten­dees on their own GC machine run an unknown sam­ple which may result in some sort of “prob­lem” in the chro­matogram. The atten­dees issue spot the prob­lem and ratio­nal­ize what is wrong, and fix it.

Hands-on Lab 10

The atten­dees with their lab part­ners go over their own data set from real cases to fig­ure out what is wrong with the atten­dees’ local laboratory.

Grad­u­ates of the group include:

STATE LAST NAME FIRST NAME   STATE LAST NAME FIRST NAME
Alaska Slone Fred New Jer­sey Her­nan­dez Steven
Ari­zona St. Louis Joe New Jer­sey Levow Evan
Cal­i­for­nia Barba Manny New Mex­ico Frechette Rod­er­ick
Cal­i­for­nia Brehmer Jeremy Okla­homa Edge Bruce
Cal­i­for­nia Ganci Eric Okla­homa Fabian Stephen
Cal­i­for­nia Gore­lick Lynn Okla­homa Hosty Tom
Cal­i­for­nia Laun­dry Vir­ginia Okla­homa Lee (x5)
Josh D.
Cal­i­for­nia Mid­dle­brook Richard Okla­homa Pat­ter­son Clint
Cal­i­for­nia Moore Ron Okla­homa Sifers Jeff
Cal­i­for­nia Sturm Craig Ore­gon Carini, Jr. Peter
Cal­i­for­nia Tie­mann Roland Penn­syl­va­nia Bar­rouk Tim
Cal­i­for­nia Was­son James Penn­syl­va­nia Man­ches­ter Brian
Cal­i­for­nia Wap­ner Terry Penn­syl­va­nia McShane (x6)
Justin
Col­orado Bussey Tim Penn­syl­va­nia Sher­man Mike
Col­orado Cessna Christo­pher Ten­nessee Garza (x2)
Mar­cos
Col­orado Her­ringer William Ten­nessee May Roger
Col­orado Savela Jason Ten­nessee McK­in­ney Rob
Col­orado Orr Rhid­ian Ten­nessee Ryan Edward
Florida Kessler Mike Texas Bal­a­gia Jaime
Florida McIn­tosh Brett Texas But­ler Jim
Geor­gia Adams Clark Texas Boatwright Nicky
Geor­gia Bab­son Rocky Texas Case Kelly
Geor­gia Caron Brian Texas Cof­fey Mimi
Geor­gia Frye Kim Texas de la Paz Brent
Geor­gia Par­man Ann Texas del Cueto Andrew
Geor­gia Stein George Texas DeLuca Matt
Illi­nois Ram­sell Don­ald Texas Flood Tyler
Illi­nois Toney Sarah Texas Grant Dean­dra
Kansas Hul­nick Les Texas Hamil­ton Stephen
Louisiana Delatte (x2)
Glynn Texas Hunter David
Louisiana Bates, Jr. James Texas McK­in­ney Troy
Mary­land Alpert Andrew Texas Mur­phy Doug
Mary­land Bruck­heim Michael Texas Ray Ben­nie
Mary­land Stamm Lenny Texas Segura Anthony
Mass­a­chu­setts Ober­hauser Gre­gory Texas Stauf­fer Phil
Michi­gan Boyle Michael Texas Trichter (x2)
Gary
Min­nesota Ram­say Charles Texas Wilder Dou­glas
Mis­souri East­man Jef­frey Utah Schatz Jason
Mis­souri Holling­shead Jeremy Vir­ginia Keefer Bob
Mis­souri Ward Carl Vir­ginia Solak– (x2)
Michael
Nebraska Dowd­ing Steve Wash­ing­ton Calla­han Linda
Nebraska Island Bell Wash­ing­ton DeBray Ted
Nevada Hayes Dale West Vir­ginia Wag­ner Harley
New Hamp­shire Russ­man Ryan Wis­con­sin Stuck­ert Lau­ren
New Hamp­shire Tenn John

There have been 89 grad­u­ates to date.

The next class (which is full) will be in April is sched­uled to have the fol­low­ing folks:

Patrick Maher

MD

Hunter Bie­der­man

TX

Wayne R. Foote

ME

John Hun­sucker

OK

Andrew Mishlove

WI

Michael J Snure

FL

Clark Adams (2nd time through)

GA

Andrew Bucher

OH

N. Cole Williams

NC

Jay M. Tiftickjian

CO

Bruce Edge (2nd time through)

OK

Brent de la Paz (2nd time through)

TX

Paul Liam McGlone

VA

Kevin Leck­er­man

PA

Jon W Woolsey

CA

Gor­don Senerius

SC

Nico La Hood

TX

Bryan E DePowell

PA

Joseph Cit­ron

GA

John j Eastland

TX

Jonathon Rands

WA

Shawn Dor­ward

PA

Jared Bartell

CA

To insure jus­tice, we need to have an edu­cated defense bar. We need more sci­en­tific pro­grams like this one.

Beer Born On Date
Beer Born On Date

Every­one wants to eat fresh food as opposed to stale food. No one wants to drink skunked beer. So, we have born-on dates on cer­tain foods and food­stuff. This allows us to judge whether or not we want to accept it and drink/eat it.

Car­rier gas purity is a big issue in Gas Chromatography. We have to remem­ber that helium is the mobile phase that car­ries the ana­lytes that are going to be ana­lyzed. It pres­sur­izes the sys­tem to “push” the injected sam­ple through the col­umn. Ide­ally, the car­rier phase is totally inert which means that it does not inter­act with the sta­tion­ary phase or the ana­lytes of inter­est. We can’t have it inter­act with either so as to change the sam­ple or the result. Although helium is not the very best car­rier gas to use per the Van Deemter equa­tions, it has become more or less the dom­i­nant car­rier gas used in foren­sic test­ing lab­o­ra­to­ries when GC is used. Helium is typ­i­cally deliv­ered in a lab­o­ra­tory and stored in a cylinder.

Again, the car­rier gas must be totally inert which means that it does not inter­act with the sta­tion­ary phase or the ana­lytes of inter­est. For test­ing of VOC’s by GC-FID, helium meets this 2 cri­te­rion. It is totally inert which means that it does not inter­act with the sta­tion­ary phase or the ana­lytes of interest.

What if this were not so?

What if the car­rier gas is not pure and does not con­tain only helium but instead con­tains things that are not inert but con­tains things that inter­act with the sta­tion­ary phase or the ana­lytes of inter­est and are even detected?

The valid­ity of our results may be compromised.

This is a real problem.

Gas man­u­fac­tur­ers place out on the mar­ket for lab­o­ra­to­ries to buy dif­fer­ent purities.

  • high purity (99.998% min. purity)
  • ultra-high purity (99.999% min. purity)
  • research grade (99.9999% min. purity)

But are you in fact buy­ing that truly that level of pure gas?

Maybe not.

The gas man­u­fac­tur­ers mea­sure purity not as it is actu­ally con­tained in the cylin­der. It is mea­sured BEFORE it gets placed into the cylin­der. The man­u­fac­turer mea­sures for oxy­gen, mois­ture, total hydro­car­bons, car­bon monox­ide, nitro­gen, and oth­ers while it is being deliv­ered to the cylin­der. Again, no one at the man­u­fac­turer tests the purity after it is actu­ally placed in the cylinder.

Cus­tomers pur­chase the gas and rent the con­tainer (This is called “demurrage”).

There will always be some level of hydro­car­bons in the cylin­der as the gas is never ever 100% pure even when it is deliv­ered for the very first time.

Every time the cylin­der is emp­tied through use, it needs to be refilled. The gas com­pa­nies sim­ply hook the cylin­der up to a vac­uum and run the vac­uum through a cycle 3 times. Helium is very, very light, there­fore, it is first in time to be removed and is very easy to remove through the vac­uum. Hydro­car­bons are much heav­ier by com­par­i­son and there­fore will be the last in order to be removed and requires more vac­uum to remove. These three cycles is sim­ply not enough to remove all the hydro­car­bons. As this process of emp­ty­ing and fill­ing hap­pens over time with the incom­plete “drain­ing” (clear­ing) of hydro­car­bons each time, hydro­car­bons will build up in increas­ing con­cen­tra­tion over time. So, it is impor­tant to note the cylin­der “born on date” to gauge the poten­tial of increased hydro­car­bons over time in the cylin­der and there­fore intro­duced into the sam­ple that is going to be analyzed.

Hydro­car­bons will inter­act with the sam­ple and the sta­tion­ary phase. Remem­ber that a Flame Ion­iza­tion Detec­tor will burn all hydro­car­bons and there­fore detect it. There­fore, it is a poten­tial source for con­t­a­m­i­na­tion and cast doubt on the valid­ity of the reported result.

Cylinder Born on Date
Cylin­der Born on Date above is 07/07 or July of 2007