New perspectives on THCA decarboxylation and accurate GC–MS
quantitation of Total THC in Cannabis using analyte protectants
Jerome Mulloor
*
, Walter B. Wilson , Lane C. Sander
Chemical Sciences Division, National Institute of Standards and Technology, 100 Bureau Drive, Gaithersburg, MD 20899, USA
ARTICLE INFO
Keywords:
Cannabis
Delta-9-tetrahydrocannabinol
Gas chromatography
Mass spectrometry
Analyte protectants
ABSTRACT
Forensic and commercial laboratories rely on well-characterized analytical methods to accurately determine
total tetrahydrocannabinol (total THC), which is the sum of decarboxylated tetrahydrocannabinolic acid (THCA)
and delta-9-tetrahydrocannabinol (Δ
9
-THC) in Cannabis sativa samples. The Agriculture Improvement Act of
2018 restricts the level of total THC allowed in Cannabis for classication as hemp for commercial purposes. Gas
chromatography with mass spectrometry detection (GC–MS) is frequently employed for Cannabis measurements
due to its simplicity and speed of analysis. However, several issues complicate the determination of total THC by
GC–MS, which are well-documented but unresolved. In the current study, the origins of potential GC–MS method
biases are investigated, and novel approaches are presented to mitigate interferences. The behavior of THCA and
Δ
9
-THC during GC–MS analyses was studied extensively using test solutions containing their isotopically labeled
analogs. A plant matrix effect was identied that signicantly increased THCA and Δ
9
-THC responses for
Cannabis extracts compared with calibrant solutions. A mechanism is proposed based on the interaction of THCA
and Δ
9
-THC with silanols present on heated inlet surfaces. The use of active site blocking agents, known as
analyte protectants, reduced these interactions and achieved suitable conditions for quantitation of total THC by
GC–MS. When Cannabis plant extracts and calibrants were processed under the recommended conditions, the
results were comparable to liquid chromatography with photodiode array (LC-PDA) analysis. The experimental
ndings ultimately provide evidence to explain the behavior of cannabinoids in the GC–MS system and offer new
options for improving the accuracy of total THC measurements.
1. Introduction
Cannabis sativa is the most commonly used recreational drug in the
United States (U.S.) [1], and its increasing prevalence signicantly im-
pacts public health, medicine, criminal justice, industry, and forensics
[2]. According to a 2022 survey, about 22 % of people (61.9 million)
over the age of 12 in the U.S. reported using Cannabis in the past year
[3]. Cannabis and its primary psychoactive constituent, delta-9-
tetrahydrocannabinol (Δ
9
-THC), have been classied as illicit
controlled substances in the U.S. since the 1970s [4]. However, legis-
lative changes over time resulted in the decriminalization or legalization
of Cannabis for medicinal and recreational purposes in many states [5].
Furthermore, the 2018 Agricultural Improvement Act (Farm Bill)
dened hemp as Cannabis containing a mass fraction of less than or
equal to 0.3 % total THC on a dry mass basis and removed hemp from the
controlled substances list [6]. Total THC refers to the sum of decar-
boxylated delta-9-tetrahydrocannabinolic acid (THCA) and Δ
9
-THC (see
Eq. 1). Consequently, as the legal landscape changed, an emerging
Cannabis industry concurrently expanded with the marketing of
cannabinoid-containing products, such as edibles, extracts, oils, and
vapes [7].
Total THC = (0.877 × THCA) + Δ
9
− THC (1)
Accurate and reliable methods for total THC analysis are indispens-
able to forensic, commercial, and compliance testing laboratories.
Forensic laboratories are responsible for providing scientic evidence to
support ndings regarding the legality of seized evidence containing
Cannabis plant material. Historically, qualitative test schemes, such as
macro- and microscopic identication of plant features and colorimetric
testing, were employed for this purpose [8,9]. After the Farm Bill was
established in 2018, forensic laboratories adapted their procedures to
measure total THC and determine whether the Cannabis sample is
marijuana (a federally controlled substance) or hemp (a legal com-
modity) according to the 0.3 % total THC threshold [10]. Furthermore,
* Corresponding author.
E-mail address: jerome.mulloor@nist.gov (J. Mulloor).
Contents lists available at ScienceDirect
Forensic Chemistry
journal homepage: www.sciencedirect.com/journal/forensic-chemistry
https://doi.org/10.1016/j.forc.2025.100668
Received 27 March 2025; Received in revised form 15 May 2025; Accepted 16 May 2025
Forensic Chemistry 44 (2025) 100668
Available online 17 May 2025
2468-1709/Published by Elsevier B.V.
