Specificity of Blood Phosphatidylethanol (PEth) as a Marker for Alcoholic Beverage Consumption
Utilizing blood phosphatidylethanol (PEth) in abstinence monitoring programs (particularly with health professional, airline pilot, and/or other professional monitoring) as a means to detect relapse to alcoholic beverage use is a relatively new development over the past decade. PEth is a minor metabolite of ethanol formed when an enzyme, phospholipase D (PLD), binds ethanol to phosphatidylcholine lipids in cell membranes, including red blood cells, in a process termed transphosphatidylation.
Although the use of PEth testing is new in alcohol monitoring, PLD has been extensively studied for decades. It turns out that the role of PLD is very complex. It has been shown that the activity of PLD is highly regulated by hormones, neurotransmitters, lipids, small monomeric GTPases and other small molecules.1 Furthermore, two major isoforms of PLD have been identified in mammalian cells, PLD1 and PLD2, each encoded by distinct genes. PLD1 activity is low until first activated by various small proteins.2 Additionally, another molecule, phosphatidylinositol 4,5-bisphosphate (PIP2), also found in cell membranes, is required as a cofactor for PLD activity. Thus, PLD activity can vary greatly between individuals. PLD is thought to play a major role in a wide range of physiological processes and diseases, including alcohol intoxication, inflammation, diabetes, phagocytosis, neuronal and cardiac signaling, and in oncogenesis.3
Other alcohol markers, such as urine ethylglucuronide (EtG) or ethylsulfate (EtS), have been shown to sometimes be present because of something extraneous (e.g. incidental or environmental ethanol exposure), from one or more of the many products containing ethanol (e.g. mouthwash, hand sanitizing gel, OTC medication, ethanol in food, etc.) rather than from alcoholic beverage consumption. Thus, a positive test does not always prove alcoholic beverage use.
Because it has been found in limited lab-based trials to require a significant amount of ethanol, >100g (about 7 standard drinks), to trigger a positive PEth test, with the typical cutoff of 20ng/ml, it has been suggested that a positive test can be considered proof of alcoholic beverage consumption.4 The rationale is that such a significant amount of ethanol to cause a positive blood PEth test is not likely to be due to extraneous sources of ethanol. The thinking is that a positive PEth test must therefore be proof of alcoholic beverage use.
However, as PEth testing has gained increasingly widespread use, numerous cases have emerged whereby individuals who have tested positive for PEth vigorously deny drinking. In some of these cases the ancillary evidence appears to support their contention. Thus, the question of false positive blood PEth testing has become a concern.
We know historically that no test is 100% specific. There have been few published studies, and fewer unpublished, that have aimed to determine the specificity of PEth for alcoholic beverage use. Existing studies have been small with few individual subjects, and all the studies have been inconclusive. So, it is important to review the facts before deciding for certain that a positive blood PEth is proof of alcoholic beverage consumption.
FACT #1
It is known that the enzyme, PLD, that catalyzes the creation of PEth from alcohol and phospholipids, has variable activity based on numerous factors mentioned above. It is logical to assume that the factors that increase PLD activity would increase blood PEth levels. This means that hormones, various disease states, other medications, etc., could affect blood PEth levels.
FACT #2
Few studies have been conducted to determine the specificity of blood PEth to detect intentional alcoholic beverage use. All existing studies, either published or unpublished, have involved very small numbers of subjects, and none have been conclusive. None have attempted to examine how medications, age, disease states, genetics, etc., might affect PEth levels. It has been suggested that such a study would need to have a significantly large group of subjects (with varying age, sex, medical conditions, etc.) carefully monitored, for example by wearing SCRAM devices or being under strict supervision, for the absence of any alcoholic beverage use while being tested for blood PEth to determine if there are false positives.5 Such a study has not been performed. Performing PEth testing in a large number of subjects who have confirmed abstinence from alcoholic beverages could help determine if there are false positive tests at various cutoff levels.
FACT #3
The half-life of PEth has been reported to range between 4-12 days. The longer half-life of PEth could conceivably lead, in some individuals, to accumulation of PEth to significant levels from one or more sources of extraneous exposure to ethanol.
FACT #4
The cutoff level for reporting a positive test is an important variable that affects specificity. Selected cutoff levels tend to be arbitrary, often without specific research to determine best cutoff levels to increase specificity without adversely affecting sensitivity. The blood PEth cutoff level used in most medical monitoring at this time is 20ng/ml. This is an arbitrary cutoff. In one study, a higher cutoff value of 221ng/ml was selected to avoid false positive readings.6 The ideal cutoff to maximize sensitivity and specificity is not clearly known.
FACT #5
When asked about false positive blood PEth tests, some experts have opined that “there has never been a confirmed false positive.” However, there have been a number of studies where groups of individuals were tested and among them were those who claimed that they had not consumed alcoholic beverages but who did test positive.7,8 Experts have opined that these subjects were likely being dishonest, falsely claiming they had not been drinking, resulting in the positive tests. In fact, there is no way to know, from the available information, if the subjects were being dishonest or not. Some, or all, of these positive tests could have been false positives.
FACT #6
A study attempted to correlate PEth levels (16:0/18.1 and 16:0/18.2) with a cutoff of 10ng/ml in 300 light social drinkers to their reported ethanol use and AUDIT-C scores.9 Only 12 subjects (4%) reported total abstinence and all were negative for blood PEth. Further analysis plotting the Receiver Operating Characteristic (ROC) curve estimated a specificity for PEth of 96.6%. If this is even close to accurate, it suggests that a small portion of individuals may have false positive readings. The authors suggest that “relevant amounts of hidden alcohol in nutrition or medication as well as inter-individual variations of enzyme activities for the formation of PEth might cause unexpected elevated concentration levels of PEth.” Even if the PEth test is 99% specific for alcoholic beverage consumption, that still means that 1 out of every 100 tests could be falsely positive. Additionally, the authors of this study determined that a blood PEth level of <112ng/ml corresponded statistically to what they called moderate drinking based on the Audit C scores (which correlated to less than an average of 10gm/d of alcohol or less than 1 standard drink per day).
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CONCLUSION
Until more extensive research is conducted, programs that monitor licensees with blood PEth tests should be careful in how the tests are used and interpreted. In particular, when the specificity of a new test is not known with certainty, a low positive test should bot be used as absolute proof of relapse. In administrative or criminal monitoring settings, this is frustrating because a test that is absolutely 100% specific is needed.
Even if not 100% specific, or if the specificity is not known with certainty, new tests can still be very meaningful and helpful. For example, if a new test, such as the blood PEth test, is positive, the monitor can question the monitoree and/or observe them more closely. When properly questioned or evaluated more thoroughly, the monitoree may admit relapse. Admission of use is the “gold standard” for diagnosing relapse. If the monitoree denies drinking and there is no other corroborating evidence of relapse, continued observation and heightened monitoring may be the best course of action. If under enhanced scrutiny the monitoree starts showing other signs of relapse (e.g. missing meetings, dishonesty, poor work performance, irritability, etc.), then testing can be intensified further and relapse will likely be detected. One certain thing about addiction is its tendency to recur. In other words, if an alcoholic relapses, it is almost certain they will continue drinking. Thus, if monitoring is continued, there will be opportunity to validate the relapse.
In conclusion, new tests for monitoring alcohol or drug use, such as the blood PEth test, can be useful even though a low positive test (<112ng/ml) by itself may not be proof of relapse. It is unlikely that the specificity of the blood PEth test will be 100%. There are essentially no tests that are perfect. Until adequate studies are performed to document more accurately the specificity of the blood PEth test, it seems reasonable that a low positive blood PEth level should not be used by itself as proof of intentional alcoholic beverage consumption. Individuals who convincingly claim they have not been drinking following positive blood PEth testing should be more carefully studied.
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1Kolesnikov YS, Nokhrina KP, Kretynin SV, Volotovski ID, Martinec J, Romanov GA, Kravets VS (January 2012). “Molecular structure of phospholipase D and regulatory mechanisms of its activity in plant and animal cells”. Biochemistry. Biokhimiia. 77 (1): 1–14.
2Peng X, Frohman MA (February 2012). “Mammalian phospholipase D physiological and pathological roles”. Acta Physiologica. 204 (2): 219–26.
3Banno Y (March 2002). “Regulation and possible role of mammalian phospholipase D in cellular functions”. Journal of Biochemistry. 131 (3): 301–6.
4Skipper GE, Thon N, Dupont RL, Baxter L, Wurst FM. Phosphatidylethanol: The Potential Role in Further Evaluating Low Positive Urinary Ethyl Glucuronide and Ethyl Sulfate Results. Alcohol Clin Exp Res. 2013 Sep;37(9):1582-6
5Hahn J, Anton R, Javors M. The Formation, Elimination, Interpretation, and Future Research Needs of Phosphatidylethanol for Research Studies and Clinical Practice. ACER. Vol 40, No 11. Nov 2016, pg 2292-2295
6Kummer N, et al. Quantification of Phosphatidylethanol 16:0/18:1, 18:1/18:1, and 16:0/16:0 in venous blood and venous and capillary dried blood spots form patients in alcohol withdrawal and control volunteers. Anal Bioanal Chem (2016) 408:825-838
7Papas RK, et al. Associations Between the Phosphatidylethanol Alcohol Biomarker and Self-Reported Alcohol Biomarker and Self-Reported alcohol Use in a Sample of HIV-Infected Outpatient Drinkers in Western Kenya. ACER (4) 8:August 2016
8Fleming MF, et al. Phosphatidylethanol Detects Moderate-to-Heavy Alcohol Use in Liver Transplant Recipients. ACER (41)4; April 2017
9Schrock A, Wurst F, Thon N, Weinmann W. Assessing phosphatidylethanol (PEth) levels reflecting different drinking habits… Drug and Alcohol Dependence 178 (2017) 80-86
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