Dr. Loralie Langman discusses the prevalence of illicit drug use in the United States and describes testing for commonly encountered drugs. She also reviews drug detection limits, confirmation cutoffs, drug metabolism, and interpretation of screening and confirmation drug testing.
Presenter: Loralie J. Langman, PhD DABCC (CC, MD, TC), DABFT
- Consultant in the Division of Clinical Biochemistry and Immunology at Mayo Clinic.
- Director of the Toxicology and Drug Monitoring Laboratory at Mayo Clinic in Rochester, Minnesota.
Welcome to Mayo Medical Laboratories' Hot Topics. These presentations provide short discussion of current topics and may be helpful to you in your practice.
Our presenter for this program is Dr. Loralie Langman, a consultant from the Division of Clinical Biochemistry & Immunology and Associate Professor of Laboratory Medicine and Pathology in the College of Medicine. Dr. Langman is also the director of the Toxicology and Drug Monitoring Laboratory at Mayo Clinic in Rochester, Minnesota.
Dr. Langman will discuss the prevalence of illicit drug use in the United States and describe testing for commonly encountered drugs of abuse. She will also review drug detection limits, confirmation cutoffs, drug metabolism, and interpretation of screening and confirmation drug testing.
I'll quickly go over the objectives. They are to understand the prevalence of drug abuse; what samples to submit for analysis; how long drugs can be detected; what tests to request; and how to interpret the results.
So let's start with a little history of drug use. Americans consume about 60% of the world's production of illegal drugs, but make up only about 5% of the world's population.
Approximately 23 million individuals in the United States use marijuana at least 4 times weekly. That’s around 8% or about 1 in 12 individuals. About 6 million people regularly use cocaine, which is about 2% or 1 in 50 individuals. And about 70% of substance abusers hold down jobs. These statistics may seem a little unusual because I think the media has really put in our minds that there are these stereotypical drug users on the street, but you can see from these statistics that is not entirely accurate.
Monitoring the Future
How do we gather some of these statistics? Well, there are organizations like Monitoring the Future. What that is, is an ongoing study of the behaviors, attitudes, and values of American secondary school students, college students, and young adults. About 50,000 students from grades 8, 10, and 12 are surveyed annually. They survey about 420 public and private schools, and they have been studying 12th graders since 1975, and 8th and 10th graders since 1991.
If we look at some of the data that’s been gathered, for 2008, if you look at the percent lifetime use of alcohol, you can see here, that the 12th graders being the golden bar, about 70% of them have used alcohol at some time in their life as compared to, let’s say, grade 8 students where it’s just shy of 40%. Now if you look at the next set of bars, that’s looking at any illicit drug use, so that combines all of the illicit drugs into 1 category, and you see by the 12th grade, just around 47% of them have used some illicit substance; whereas, if you look at 8th graders, it’s just about 19%.
If you take that graph and break it out a little bit, the 2 sets of first bars are alcohol and illicit drug use again, but you break out the illicit drugs, you have marijuana as the most highly prevalent of those illicit drugs, with marijuana use being just over 40% in 12th graders. As you go down the list, you have amphetamine-type stimulants, cocaine, LSD, anabolic steroids, and heroin.
Now if you break out the daily use category, what I've done here is shown you 8th, 10th, and 12th graders broken out as individuals comparing alcohol and marijuana use, alcohol being the blue bar and marijuana being the purple. Now remind you, this is daily use, and if you focus on the 12th graders, about 2.8% of them use alcohol daily, whereas, what's interesting is about 5% use marijuana daily.
Ethanol - Annual Use
If you look at these statistics over time, going back to 1991, you have the 12th graders again being the gold bars, and the 10th pink and the 8th the blue, you can see that the alcohol use over time has decreased slightly in those categories, but it’s not statistically significantly different.
Any Illicit Drug - Annual Use
Comparing that to illicit drug use, you can see it’s changed quite a bit over time. If you compare it to the early 1990s, the actual prevalence of illicit drug use has gone up in just about all of the different grade levels.
Now this is a different category. This is inhalants, so these are things like gasoline, glue, paint thinner, those kinds of things, and you can see here in annual use, the prevalence is a little bit different where you have the 12th graders being much lower than the 8th graders. This is primarily due to the ease of access for these drugs. It’s much easier for a younger person to get a hold of something like, for example, glue; whereas, a 12th grader has easier access to some of the other abuse substances.
So let’s shift gears a little bit and talk about drug testing itself. Why would we want to test for drugs? Well there are 3 reasons right here. One is to confirm a diagnosis, to rule-in or rule-out drug use in an individual patient. Another reason might be to determine the extent of intoxication of the patient who is sitting in front of you as you’re performing the examination. And another reason might be regulatory; for example, a workplace drug testing program. This list is by no means inclusive, but it is probably the most common reasons that people use drug testing.
When you do, do drug testing you must ask yourself a couple of questions. What are you trying to detect? Not only does that fall into what type of drug are you trying to detect, but which of the reasons you are going to be detecting drugs and what are you trying to accomplish? What is it that is making you want to test drugs in this individual?
Well, going along with that, you have to decide what specimen to collect. The question you have to ask yourself now is, is the person I am examining right now under the influence of drugs? If that is the question, then blood or serum is the most appropriate. To decide if that person is impaired right now, that’s the best one. However, if the question you’re asking is was an individual exposed to a drug or using the drug in the past, then urine would be a more appropriate sample, as it goes and gives you a longer time detection window, which will be discussed later on.
In the Lab
So how do we do it in the lab?
Drugs of Abuse
Well, we basically do it in two stages. We have stage one, which we call a screening procedure, and stage two, which is confirmation, which can also be called an identification step.
In the screening step, if it’s done in the laboratory, it’s commonly an immunoassay. And here are some common methods that we use to detect them. The actual mechanism of these assays will not be discussed in this presentation.
Point of Care
However, there are also point-of-care devices. Again, some of them are immunoassay based, but some of them are color based. In other words, you add 2 chemicals together, for example, and get a blue color—is the test for acetaminophen.
How do they Work?
Like I mentioned, most of the testing for screening is an immunoassay-based method. Well, how do they work?
Identify Negative Samples
Well, in the very simplest form, they detect compounds that look like the drug you're interested in. Why do I say that? Well, it's actually interesting to think of it, but the main purpose is actually to identify negative samples.
Sensitivity vs Specificity
So what we have here is a balance between sensitivity and specificity of analytical methods. We want to design the drug screening assay so that the number of false-negatives is very low, but because of the balancing act, it does tend to make the number of false-positives a little bit higher.
Now false-positives and some false-negatives can be attributed to a phenomenon called cross-reactivity. Remember, the antibody is looking for substances that look like the drug, and we measure the degree or assess the degree of cross reactivity by a percentage.
Here's an example. Oxycodone, let’s say, is 5% cross-reactive with the opiate immunoassay. Well, what does that mean? It means you need 20 times as much oxycodone to give you a positive result in that screen, which is about 6000 ng/mL of oxycodone if you’re using a 300 ng/mL cutoff.
Now when we have drug testing, the top 4 requests that a laboratory typically gets are cannabinoids, which is marijuana, cocaine and its metabolite, amphetamine-type stimulants, and opiates. Now when we say opiates, the assays really look for morphine and codeine, and that's important to remember.
Antibody- or Immunoassay-based
Now again, here are the top 4 that are requested, but it is certainly not the only list that can be detected. We can also look for PCP, barbiturates, benzodiazepines, methadone, ethanol, propoxyphene, methaqualone, and LSD. Again, it is not an inclusive list, but these are the more common. Now it's also important to remember that all of these are screens, since most of them are antibody- or immunoassay-based, ethanol being the exception because it’s an enzymatic-based assay.
Identify the Positive Result
So now that we have a screening method, how do we actually identify what's causing that positive result.
Well, that's where the confirmation testing or the identification comes in. It is almost always done in the laboratory. There aren't really any point-of-care devices that will satisfy the requirements. Typically, it's a gas chromatography-mass spectrometry or liquid chromatography-mass spectrometry or tandem-mass spectrometry, but the thing that's important to remember is it identifies the drug with certainty. If that type of testing reports amphetamine, for example, that drug has been identified.
Now, of course, drugs of abuse are not the only drugs that we want to be able to test for. So there are other testing methods that we can use or testing devices, one of which is a test called a prescription or over-the-counter drug screen. This could go by other names, but basically it’s a description of what it can detect. Again, it will not detect drugs of abuse; it’s basically for those prescription and over the counter drugs. Again, it is done in the laboratory by the same types of testing methodologies; however, the sample preparation is different and, therefore, why you can’t detect the same drugs as you can for the drugs of abuse.
Now the other thing to remember is drug testing. Well, we try to look for as many drugs as possible, but we’re never going to find every drug that’s out there. Imagine that we’re fishing and we’re casting a net and we’re scooping out all those fish and those fish are drugs. You know that some of those fish are going to sneak through the holes in the net. But sometimes we can only detect fish that are at lots and lots of numbers of them. In other words, we can only detect drugs at concentrations that are probably above what’s seen therapeutically. So it’s very useful in the evaluation of a patient with an overdose, but not necessarily always useful when you’re looking for therapeutic compliance. That’s when the request for a specific drug is very important and probably more useful than to go for a broad-base drug screening method.
Drugs Not Commonly Tested
Now here are some examples of those fish that sneak through the holes in the net. In other words, drugs not commonly tested. They are listed here. Ones that are important to remember are anabolic steroids; chloryl hydrate; methylphenidate; diuretics; GHB, which is gamma hydroxybutyrate; ketamine; the laxatives; most, but not all, of the nonsteroidal antiinflammatory drugs; inhalants; and fentanyl. But, that is again not an inclusive list.
Detected But Not Reported
Now sometimes we detect drugs but don’t report them and drugs that fall into that category tend be things like caffeine, which is present in so many substances; it’s found in so many people that to report it every time would not be useful. However, should it be found in very, very high potentially dangerous levels, it would be reported. Another drug would be nicotine and its metabolite cotinine.
But it's also important to know what test menu is available and, of course, the corollary, what is not. Really important to find out if we can detect a drug is to give us a call. Check with the laboratory.
Now it’s important to be able to interpret drug screens; need to know a little bit about drug metabolism. Basically it’s divided into 2 phases: Phase 1 and Phase 2. Phase I being comprised of oxidation and reduction of the drugs, usually mediated by something like an enzyme of the cytochrome p450 class, but again there are other drugs. The phase 2 reactions are conjugation reactions and can conjugate the drugs with the list of compounds indicated here. Generally, the purpose of the metabolism is to generate compounds that are more soluble and that helps aid in excretion.
Now I'm going to quickly go over the metabolism of some of these drugs. First I'm going to start with morphine. Morphine, as you know, is an analgesic. It's metabolized by UGT2b7 and is a phase 2 reaction as its glucuronidated. One of the metabolites, which is morphine-6-glucuronide, is a small percentage about 5% to 15% but is actually an active metabolite. The major metabolite is the morphine-3-glucuronide, which is not inactive and, of course, you can glucuronide both the 3 and the 6 making it a digluconuride metabolite.
Codeine is an interesting drug given as an analgesic, but codeine has its analgesic property not from codeine, but from its demethylated metabolite, morphine. Morphine we've already discussed on the previous page is glucuronidated and excreted.
Now heroin in this country is an illicit drug, although it is one of the most potent analgesic medications that’s available today. Heroin, or diacetyl morphine, the first metabolic step is a hydrolysis to form monacetyl morphine. This process is very rapid and it is very, very infrequent that we actually find heroin present, but its unique characteristic metabolite, 6-acetyl-morphine we can detect. 6-acetyl-morphine is then hydrolyzed again to morphine, which, of course, we also can detect in the laboratory and is glucuronidated and then excreted in the urine.
Again, oxymorphone, another drug that is given as an analgesic is simply glucuronidated and excreted much like morphine.
Oxycodone is then also metabolized to oxymorphone, which is then glucuronidated and excreted.
Hydromorphone, glucuronidated and excreted.
And hydrocodone metabolized to hydromorphone, which is glucuronidated and excreted. So you can see that a lot of the opiate analgesics have what we call active metabolites.
Now we're going to move on to another opioid, which is methadone. Methadone has numerous metabolic reactions, but the one that we measure in the laboratory is EDDP. The reason we measure EDDP is it is a characteristic metabolite such that its presence indicates that methadone has, in fact, been consumed and the methadone has not been exogenously added to the urine sample.
Now cocaine undergoes numerous metabolic pathways, but the one that the laboratory commonly measures it the conversion of cocaine to benzoylecgonine. So you will see that both cocaine and benzoylecgonine are detected and reported in the assay.
THC, otherwise known as tetrahydrocannabinol, the active component of marijuana, undergoes numerous oxidation steps to the compound that we measure in the laboratory, carboxy THC. This is not the final metabolic pathway of THC and actually undergoes another set of numerous reactions to the metabolites indicated on the slide.
Now in the amphetamine-type stimulants category, amphetamine being the prototype of course, but methamphetamine; MDMA, which is ecstasy; and MDA, which is methylenedioxyamphetamine; also fall into the category of amphetamine-type stimulants. Now all of these have the potential to be drugs that would be used as individual drugs. However, it's also important to remember that methamphetamine is demethylated to amphetamine, so another example of an active metabolite, and MDMA is demethylated to MDA, again another example of active metabolites.
Now benzodiazepines are a very complex group of drugs. There is somewhere around 32 prescription benzodiazepines that are available in the United States. Diazepam, which is pictured here, is sort of the prototype of those, and the basic structure is similar to this with variations as you'll see in some of the slides to follow.
The metabolism of some of the benzodiazepines is very simple. Take lorazepam for example; it's glucuronidated and excreted.
If you look at clonazepam and flunitrazepam, which actually flunitrazepam is not available as a prescription drug in the United States but also goes by the name of Rohypnol, is frequently associated with drug-facilitated sexual assault. Both of these are metabolized to 7-amino metabolites, which are then glucuronidated and excreted.
Triazolam, alprazolam, midazolam, and flurazepam all are hydroxylated into their respective metabolites, which are glucuronidated and excreted.
Now, I had kind of alluded to the fact that benzodiazepine metabolism was complex and here is what I mean. Diazepam, which of course, is a prescription drug is metabolized to nordiazepam, which is also a prescription drug, and nordiazepam is then metabolized to oxazepam, which is also a prescription drug. Now, diazepam also has another pathway, which is metabolized to temazepam, which then again is metabolized to oxazepam. And just when you thought it was simple, both temazepam and oxazepam are glucuronidated and excreted. Now another drug, chlordiazepoxide, also ultimately ends up being metabolized to nordiazepam.
So here's where it gets interesting. If you detect oxazepam in the urine of a patient, they could be taking oxazepam, they could be taking nordiazepam, they could be taking diazepam, or they could be taking chlordiazepoxide. It is very difficult to tell which one of those drugs was the actual drug given to the individual. Occasionally, you will find multiple metabolites. For example, you may find nordiazepam, temazepam, and oxazepam, strongly suggesting diazepam use, but still cannot rule out that nordiazepam was also administered. Nor can you rule out that oxazepam was also administered.
Now one of the most frequently asked questions in the laboratory is how long can you detect a drug? Well, of course, it varies from drug to drug, but it also varies from sample to sample. Here is an example looking at cocaine and its metabolite. In blood, you can probably only find evidence of cocaine use up to 24 hours, but only through its metabolite benzoylecgonine. If you look in urine, you can find it for up to 3 days.
And for the rest of this discussion on detection times, we will be focusing on urine drug testing mostly because it is the most frequently used type of sample for the assessment of drug use.
Now another thing to remember is how long we can detect a drug depends on how low we can detect our drug; in other words, what is our detection limit?
Drug Detection Limits
Now you’ll see frequently the term cutoff. And the reason we use the term cutoff is because a positive result is reported if the concentration measured is above that prespecified value or cutoff.
Drug Detection Limits: Opiates
Now if we use here these tables looking at the opiates, we have a screening cutoff of 300 ng/mL, and the confirmation cutoff of 300 ng/mL would be an example of what I’m referring to.
Drug Detection Limits: Amphetamines
So for amphetamines, we have a screening cutoff of 1000 ng/mL and a confirmation cutoff of 500 ng/mL. Now the thing that’s very interesting, is if you notice the double asterisk note with methamphetamine, methamphetamine has a concentration cutoff of 500 ng/mL, but amphetamine must also be present for that to be reported.
Drug Detection Limits: Cannabinoids/Cocaine
Now we’ll look at some of the other drugs. Cannabinoids, with the screening and confirmation cutoff, as indicated, at 50 ng/mL and 15 ng/mL; cocaine, where we have a screening cutoff of 300 ng/mL and a confirmation cutoff of 150, but we’re actually confirming its metabolite, benzoylecgonine; and PCP at the screening/confirmation cutoff of 25 ng/mL.
Now that we have those screening cutoffs, based on those detection limits, the time that we can detect drugs are as follows.
Detection Times: Stimulants
For the stimulant drugs including amphetamine, methamphetamine, some of the amphetamine variants including MDA and MDMA (remember ecstasy), cocaine and methylphenidate (or Ritalin), there usually, and you can see there, but it usually averages about 1 to 3 days.
Detection Times: Opioids and Morphine Derivatives
If you look at the opioids and the morphine derivatives: morphine; the heroin metabolite 6-acetyl-morphine (being less than 24 hours); but the morphine, codeine, oxycodone are about 1 to 3 days. And that also applies to a lot of the other opiates including hydrocodone and hydromorphone. Methadone and its metabolite EDDP, we can see from 1 day to 1 week. Now that sounds like a very long window of time, but it’s basically because the metabolic half-life of that drug is extremely variable from individual to individual, so it’s very difficult to predict.
Detection Times: Depressants
Now if you look at the depressant drugs, we have alcohol, which we can detect in urine for 6 to 10 hours; very, very dependent, of course, on the amount of alcohol that was consumed. The barbiturates have different detection times depending on their activity. Short-acting barbiturates tend to have a short half-life and, therefore, are detected for shorter periods of time.
Similarly, the benzodiazepines; the very short-acting benzodiazepines, can only be detected for up to a day. Drugs like methaqualone could be detected for up to 2 weeks because of a much longer half-life.
Detection Times: Hallucinogens
Now if you look at the hallucinogen drugs, LSD for example, can be detected for 8 hours; again a very short half-life. Mescaline, PCP, and Psilocybin (or magic mushrooms) can be detected for varying periods of time as well. Now you’ll also notice that I have the amphetamine variants here, listing again MDA and MDMA. The reason it’s been listed twice is not because the half-life or the detection window is different, but because those drugs also have hallucinogenic properties.
Marijuana, of course, is probably the drug that most people are interested in simply because it’s an illicit drug of abuse that’s used the most frequently. Unfortunately, its detection period is hugely variable based on a variety of different factors.
One is the frequency of use; a single use versus somebody who uses the drug daily at very large amounts. If you have somebody who uses the drug for 1 time, you might be able to find it for 3 days. Conversely, if you have somebody who has been smoking marijuana for a long period of time, has been using it multiple times a day, and happens to have a high body mass index, the drug can be detected from approximately 3 to 6 weeks. Why did I mention body mass index? The drug itself is very lipophilic and once it’s absorbed into the fat cells, it takes a very long time for it to be removed from those fat cells and, thus, extends the period of detection.
Now, it’s also important to remember that since we can detect a drug in urine for days or weeks, a positive result doesn’t necessarily mean that the patient is under the influence of the drug at the time the sample was collected. And that is very important when you’re screening patients in the emergency room.
Now we’re going to get into a little bit of the nitty gritty here of drug screen interpretation. When a drug screen is positive, and the confirmation test is positive, it pretty much is obvious that the drug indicated in the report was detected at or above the cutoff value that is also indicated in the report. That’s probably the easiest one to figure out. The individual was exposed to the drug.
But, remember, it doesn’t tell you when.
So here’s another common circumstance where the drug screen and the confirmation are both negative. It could mean the drug was not ingested. It could also mean the drug was ingested too far in the past to be detectable. An example would be LSD. The detection window for LSD is only about 8 hours, but if the request for testing was performed 3 days following LSD use, we won’t be able to find it. It could be the drug was ingested, but it was below those concentration cutoffs and, therefore, unable to be reported. And it could also be that the drug ingested was not included in the screen. For example, the patient may have taken pseudoephedrine; well, that is not an illicit drug and is not included in the illicit drug of abuse screen.
Now a not uncommon circumstance is when the drug screen itself is positive, but the confirmation test is negative. It could be that no drugs were ingested at all and what you have is a cross-reaction with the antibody and it’s just found something else.
It could be that the drug is ingested in the very remote past, such that metabolites are cross-reacting with the antibody causing the positive screen, but those metabolites are not detected in the confirmatory method.
It could also be again that drugs were detected at concentrations that were below the reportable confirmation cutoff.
And again, it could be that drugs ingested were not included in the confirmation assay. An example would be when you have the amphetamine screen coming up positive and it was due to pseudoephedrine, but because pseudoephedrine was not detected in the confirmation assay, it was not reported.
So the next question then becomes, well what do we actually include in the confirmation assay? Screening assays frequently look for a class of drugs. I will look at say, for example, benzodiazepines, but the confirmation assay doesn’t necessarily include all of the drugs and frequently only looks for illicit drugs. The best way to find out what the lab includes in their confirmation assay is give us a call.
So very quickly to summarize the interpretation, a positive drug screen tells you something. It tells you that the patient has drug X in their system.
But don’t forget context: the patient’s drug history, drug metabolism, and timing of collection. Remember the benzos and how complicated their drug metabolism was?
What about a negative drug screen? Well, a negative drug screen does not necessarily rule-out drug use. It could be again drugs were not ingested, they were ingested too far in the past, they were below the cutoff concentrations for reportable, and the drugs ingested are not included in the screens performed.
Know what the lab can and cannot detect, but more importantly, let us know what you need. Good drug screen interpretation requires 2-way communication. Give the lab director a call. That’s why I’m here.