Drug and Alcohol Dependence 227 (2021) 108962

Contents lists available at ScienceDirect

Drug and Alcohol Dependence

journal homepage: www.elsevier.com/locate/drugalcdep

Prevalence of new psychoactive substances (NPS) in Brazil based on oral

fluid analysis of samples collected at electronic music festivals and parties
Kelly Francisco da Cunha a, b, Karina Diniz Oliveira a, b, Marilia Santoro Cardoso a, b,
Ana Carolina Furiozo Arantes a, b, Pedro Henrique Piras Coser b, Lucas de Noronha Lima b,
Ana Cristhina Sampaio Maluf a, b, d, Maria Angélica de Castro Comis d, Marilyn A. Huestis e,
Jose Luiz Costa a, c, *
a
Campinas Poison Control Center, University of Campinas (UNICAMP), Campinas, SP, 13083-859, Brazil
b
Faculty of Medical Sciences, University of Campinas (UNICAMP), Campinas, SP, 13083-859, Brazil
c
Faculty of Pharmaceutical Sciences, University of Campinas (UNICAMP), Campinas, SP, 13083-859, Brazil
d
ResPire Harm Reduction Project, Centro de Convivência É de Lei, São Paulo, SP, 01019-020, Brazil
e
Institute of Emerging Health Professions, Thomas Jefferson University, Philadelphia, PA, USA

A R T I C L E I N F O A B S T R A C T

Keywords: Background: New psychoactive substances (NPS) use is a worldwide public health issue. Knowing the prevalence
Oral fluid of NPS guides public health and legal policies to address the problem. The objective of this study was to identify
New psychoactive substances (NPS) NPS in Brazil through the analysis of oral fluid (OF) samples collected at parties and electronic music festivals.
Brazil
Methods: Anonymous questionnaires and oral fluid samples were collected from volunteers (≥18 years) who
Electronic music festival
MDMA
reported the consumption of at least one illicit psychoactive substance in the last 24 h. Oral fluid sample col­
Ketamine lections occurred at eleven parties and two electronic music festivals over 16 months (2018–2020). Question­
naire answers were matched to oral fluid toxicological results.
Results: Of 462 oral fluid samples, 39.2 % were positive for at least one NPS by liquid chromatography‒tandem
mass spectrometry (LC–MS/MS). The most prevalent NPS was ketamine (29.4 %), followed by methylone (6.1 %)
and N-ethylpentylone (4.1 %); however, MDMA was the most commonly identified (88.5 %) illicit psychoactive
substance. More than one drug was identified in 79.9 % of samples, with two (34.2 %) and three (23.4 %)
substances most commonly observed. Only 5 % of volunteers reported recent NPS consumption.
Conclusion: MDMA is still the most common party and electronic music festival drug, although NPS were iden­
tified in more than one-third of oral fluid samples.

1. Introduction
New Psychoactive Substances (NPS) mimic the effects of conven­
tional drugs of abuse but may not be prohibited by international drug
laws (DEA, 2019). NPS are a worldwide public health problem (UNODC,
2020a), with many intoxications and deaths (Adams et al., 2017; Pichini
et al., 2018; Zaami et al., 2018; Zawilska et al., 2020). A major risk of
inadvertent consumption occurs when conventional drugs are adulter­
ated with NPS, such as the addition of fentanyl and its analogues to
heroin, methamphetamine or cocaine samples, or when there is the
complete substitution of a conventional drug, e.g., oxycodone, without
the users’ knowledge (Misailidi et al., 2018).
In 2018, the European Monitoring Centre for Drugs and Drug
Addiction (EMCDDA) reported, on average, one new NPS per week, with
current monitoring of more than 730 different substances. Although the
number of newly introduced NPS decreased compared to 2013–2014,
there is still a considerable number of new substances emerging in the
illicit market. Currently, the classes with the highest incidence of new
drugs are opioids and benzodiazepines. Although NPS consumption is
less than for conventional drugs of abuse, two thirds of European
countries are concerned about intake of these substances by vulnerable
and high-risk individuals (EMCDDA, 2019).
NPS consumption varies worldwide. In the Netherlands, the most
identified classes between 2013–2017 were phenylethylamines and
synthetic cathinones (Hondebrink et al., 2020). NPS poison control
center reports from health professionals increased from 4 to 11 % in the

* Corresponding author at: Campinas Poison Control Center, University of Campinas (UNICAMP), Campinas, SP, 13083-859, Brazil.
E-mail address: jose.jlc@fcf.unicamp.br (J.L. Costa).

https://doi.org/10.1016/j.drugalcdep.2021.108962
Received 19 February 2021; Received in revised form 15 June 2021; Accepted 10 July 2021
Available online 12 August 2021
0376-8716/© 2021 Elsevier B.V. All rights reserved.
K.F. da Cunha et al.
same period, with at least 50 % related to phenylethylamines.
Conversely, in the USA, the National Center for Health Statistics, linked
to the Centers for Disease Control and Prevention (CDC), showed a
significant increase in deaths due to synthetic opioids (except metha­
done) from 2013 to 2018 (Hedegaard et al., 2020).
Many countries monitor NPS emergence and notify first responders,
medical personnel and public health officials to address the problem.
The National Forensic Laboratory Information System (NFLIS) (2021) of
the US Drug Enforcement Administration, NPS discovery of the Center
for Forensic Science Research and Education (2021), the Early Warning
Advisory on NPS of the United Nations Office on Drugs and Crime
(UNODC, 2021), and the EU Early Warning System on NPS of the
EMCDDA (EMCDDA, 2021a), in Europe, are examples of these initia­
tives. Brazil does not yet have an early warning system. Consumption of
psychoactive substances is common in bars and music festivals
(EMCDDA, 2019; Fernández-Calderón et al., 2018; Palamar and Keyes,
2020), with testing panels of psychoactive substances in Europe
including most commonly 3,4-methylenedioxymethamphetamine
(MDMA), cocaine and amphetamines (EMCDDA, 2019).
The NPS term was designated for substances not controlled by the
1961 Single Convention on Narcotic Drugs and the 1971 Convention on
Psychotropic Substances (UNODC, 2021) by the United Nations Office
on Drugs and Crime (UNODC). Due to the dynamics of the illicit market
and the inherent risk to public health, some substances reported in this
study were controlled under these conventions and would not be
considered as NPS (e.g. mephedrone, 25I-NBOMe). However, for
epidemiological purposes, to facilitate presentation and understanding
the data, and considering the importance of this public health issue,
maintaining the initial classification as NPS was more appropriate. Ke­
tamine was also designated as NPS following the UNODC NPS categories
(UNODC, 2021).
Oral fluid (OF) is currently the main alternative matrix to urine for
monitoring drugs and is well accepted by volunteers. It provides a non-
invasive, observed, and gender-neutral collection (Desrosiers and
Huestis, 2019). Festival attendees (n = 396) in Miami, FL, USA provided
126 blood, 226 urine and/or 374 OF samples (Mohr et al., 2018).
Compared to other alternative matrices such as hair and urine, OF also
offers the advantage of better identifying recent drug consumption. Oral
fluid more likely contains the parent drug, facilitating NPS identification
compared to urine samples. The lack of NPS metabolism studies makes it
difficult to identify the appropriate metabolites. Although stability may
be a concern for some analytes (e.g., benzodiazepines, synthetic cath­
inones), addition of the elution buffer facilitates recovery and stability
but also dilutes the sample requiring highly sensitive analytical instru­
mentation. In addition, NPS and MDMA OF concentrations correlate
better with blood concentrations than those in urine despite initially
higher OF concentrations (Mohr et al., 2018; Desrosiers and Huestis,
2019).
The objective of this study was to identify the consumption of NPS
and heroin, cocaine and methamphetamine in Brazil through the anal­
ysis of oral fluid samples collected at parties and electronic music
festivals.
2. Materials and methods
2.1. Chemicals
All psychoactive standards were obtained from Cayman Chemical
(Ann Arbor, MI, USA) or Cerilliant (Round Rock, TX, USA). Methanol
and formic acid (98–100 %) were obtained from Merck (Darmstadt,
Germany) and Scharlau (Barcelona, Spain), respectively. Methyl t-butyl
ether (MTBE) was purchased from Sigma-Aldrich (St. Louis, MO, USA)
and sodium tetraborate from L.S. Chemicals (Ribeirao Preto, SP, Brazil).
Ultrapure deionized water was supplied by a Milli-Q RG unit from
Millipore (Billerica, MA, USA). All solvents employed in the extraction
were HPLC grade.
2
Drug and Alcohol Dependence 227 (2021) 108962
2.2. Sample collection, transport, and storage
Research was conducted in accordance with the ethical standards of
the University of Campinas. Copies of the informed consent form were
available to volunteers at all events.
OF sample collection occurred at 13 different events from September
2018 to January 2020. Eleven events were parties with an average
duration of 8 ± 2 h and 735 ± 350 participants, and the other two events
were electronic music festivals with more than 5000 participants over
6–8 days. OF collection utilized the Immunalysis Quantisal™ device
(Pomona, CA, USA) and occurred in the harm reduction services area of
electronic music festivals. Prior authorization from each event’s orga­
nizing committee was received. Volunteers were ≥18 years old, received
information about the research objectives, completed an anonymous
and confidential questionnaire, and donated OF according to the man­
ufacturer’s instructions (Immunalysis, 2020). The oral fluid collection
pad was placed under the tongue and kept until the volume adequacy
indicator turned blue or for 5 min. The collection pad was placed into
the tube containing the elution buffer, capped and mixed. Six questions
documented the age, gender, sexual orientation, education, estimated
time of last consumption of any illicit psychoactive substance, and
estimated time of last consumption of illicit psychoactive substances in
the last 24 h. The questionnaire and OF sample were paired and
numerically identified, allowing volunteers to anonymously obtain re­
sults of the OF toxicological analysis seven days after the end of the
event by individual sample/questionnaire number and a personal
password.
Quantisal™ OF collection devices were stored in a refrigerator at 4
◦
C before transport to the laboratory on dry ice. During collection at
festivals over several days, samples were stored in a refrigerator at 4 ◦ C
before transport to the laboratory. Five hundred microliters of the OF-
elution buffer mixture was aliquoted into polypropylene tubes for
extraction, with the remaining volume aliquoted into two cryotubes and
stored at − 20 ◦ C.
2.3. Sample extraction and instrumentation
Sample preparation was performed with 500 μL of the OF-elution
buffer mixture, 25 μL deuterated internal standard mixture (THC-D3
50 ng/mL, diazepam-D5 50 ng/mL, MDMA-D5 20 ng/mL, LSD-D3 20 ng/
mL, fentanyl-D5 200 ng/mL and morphine-D5 200 ng/mL) and 500 μL
saturated sodium tetraborate aqueous solution, followed by a liquid-
liquid extraction with 2 mL MTBE. After vortexing for 2 min and
centrifugation (4500 rpm/5 min), 1.7 mL organic phase was evaporated
under nitrogen (40 ◦ C, 5 psi) and reconstituted with 100 μL methanol.
Two microliters of the extract was injected onto the Nexera UHPLC
chromatographic system coupled to a LCMS8060 triple quadrupole mass
spectrometer (Shimadzu, Kyoto, Japan). Validated limits of detection (n
= 18) were 0.05, 0.1 or 1 ng/mL for all analytes, except 5 ng/mL for
acetyl norfentanyl and mescaline. Detailed information regarding
chromatographic conditions, MRM optimized for NPS and drugs of
abuse substances and qualitative validation results were previously
published in da Cunha et al. (2020). A complete list of all analytes
included in the LC–MS/MS method and their limits of detection are
shown in Table 1.
A quality control sample prepared with authentic blank oral fluid
fortified with NPS-working standard solution to 10 ng/mL, and a
negative authentic oral fluid sample were included in each sample
batch.
2.4. Analysis of results
LC–MS/MS results are presented as percent positivity for each sub­
stance in the total sample and based on type of event (party or electronic
music festival). Volunteers’ questionnaire answers were compared to
their toxicological results. Statistical analyses were performed using the
K.F. da Cunha et al. Drug and Alcohol Dependence 227 (2021) 108962

Table 1 metab metabolite.

Psychoactive substances and metabolites and their limits of detection investi­
gated in the oral fluid by liquid-liquid extraction and LC–MS/MS analysis.
GraphPad Prism software (version 5.0).
Analytes and limits of detection (ng/mL)
Access to sample results on the website was monitored for the last
NPS Fentanyl Acetyl fentanyl (0.05), acetyl norfentanyl five events, three parties and two electronic music festivals to assess
analogues (5, metab), acrylfentanyl (0.05), alfentanil whether participants were interested in checking their own consump­
(0.1), carfentanil (0.05), fentanyl (0.05),
tion after a laboratory toxicological analysis.
furanylfentanyl (0.05), norfentanyl (1,
metab), remifentanil (0.1), sufentanil (0.1),
thiofentanyl (0.05), valerylfentanyl (0.05) 3. Results
Synthetic AB-FUBINACA (0.1), AB-PINACA (0.05),
cannabinoids APINACA (0.05), AM2201 (0.05), AM2233
Over 16 months, 462 OF samples were collected at 13 different
(1), HU-211 (1), JWH-015 (0.05), JWH-019
(0.05), JWH-073 (0.05), JWH-081 (0.05), events, 231 at parties and 231 at two electronic music festivals. The
JWH-122 (0.05), JWH-175 (0.05), JWH- mean age of participants was 25 (18–51, median 23) years; 55.4 % were
200 (0.05), JWH-203 (1), JWH-210 (0.05), males, 64.5 % reported being heterosexual. About 59 % were under­
JWH-250 (1), JWH-176 (1), RCS-4 (0.05), graduate students, and 32.3 % had a higher education or graduate de­
RCS-8 (0.05), XLR11 (0.1), PB22 (0.05),
gree. Most (88.5 %) reported consumption of an illicit psychoactive
MMB-FUBINACA (0.05)
Synthetic Cathinone (1), methcathinone (1), butylone substance in the last three months. Detailed demographic data are
cathinones (0.05), pentylone (0.05), N-ethylpentylone shown in Supplementary Table 1.
(0.05), mephedrone (0.05), methedrone MDMA was detected in 88.5 % of OF samples, followed by Δ9-
(1), pentedrone (1), benzedrone (0.05), 4-
tetrahydrocannabinol (THC) in 73.6 % (Supplementary Table 2). A total
chloroethcathinone (0.05), dipentylone
(0.1), dimethylone (1), MDPV (0.05),
of 181 samples contained at least one NPS (39.2 %), with ketamine (75.1
ethylone (0.05), methylone (0.05), alpha- %) the most prevalent (Fig. 1), followed by methylone (15.5 %) and N-
PVP (0.05), pyrovalerone (0.05), ethylpentylone (10.5 %). In addition to these substances, other synthetic
naphyrone (0.05), 4-fluoromethcathinone cathinones included dipentylone (5.0 %), eutylone (2.2 %), mephedrone
(1), eutylone (0.1)
(1.1 %), MDPV (0.6 %), and 4-chloroethcathinone (0.6 %). Hallucino­
Phenethylamines 25B-NBOH (0.1), 25B-NBOMe (0.1), 25C-
NBOH (0.1), 25C-NBOMe (0.05), 25D- genic phenethylamines 25I-NBOH (3.9 %), 25C-NBOH (2.2 %) and 25B-
NBOMe (0.1), 25E-NBOH (0.05), 25E- NBOH (0.6 %) were also identified, primarily in combination with 25I-
NBOMe (0.1), 25G-NBOMe (0.1), 25H- NBOH, 2C-X (0.6 %), and 25I-NBOMe (0.6 %). The tryptamine deriva­
NBOMe (0.05), 25I-NBOH (0.05), 25I-
tive 5-MeO-MiPT (1.7 %) was detected in three festival samples and
NBOMe (0.05), 25N-NBOMe (0.05), 25T2-
NBOMe (0.05), 2C-B (1), 2C-C (1), 2C-D
piperazine mCPP (m-chlorophenylpiperazine, 0.6 %) in one festival
(0.1), 2C-E (1), 2C-G (1), 2C-I (0.1), 2C-T sample. No synthetic cannabinoid included in the analytical method (N
(1), 2C-T-2 (1), 2C-T-4 (0.1) = 22) was detected.
Other NPS LSD (0.05), amphetamine (1), Psychoactive substances identified in OF from parties and festivals
and drugs methamphetamine (1), MDA (1), MDMA
were similar (Supplementary Table 2); however, ketamine was the
of abuse (1), MDEA (1), PMA (1), 4-MTA (1), DOET
(1), DOM (1), 3-methoxi PCP (1), BZP (1), fourth most prevalent substance in samples collected at electronic music
diethylpropion (0.1), DMT (0.1), DOB (1), festivals (45.9 %), and the sixth most prevalent at parties (13.0 %).
2-MAPB (1), 5-MAPB (0.05), ketamine Among the NPS-positive OF samples, eutylone, 5-MeO-MiPT, 4-chlor­
(0.1), norketamine (1, metab), etizolam (1), oethcathinone, MDPV, and mCPP were only detected at festivals,
TFMPP (0.1), OH-LSD (1, metab), 5-MeO-
MiPT (0.05), mCPP (1), fenproporex (1),
whereas 2C-X, mephedrone, 25B-NBOH, and 25I-NBOMe were only
mescaline (5), THC (1), methylphenidate detected at parties. But ketamine, methylone, and N-ethylpentylone
(0.05), ephedrine (1), cocaine (5), were the most prevalent NPS in both locations.
benzoylecgonine (5, metab), cocaethylene Ingestion of multiple illicit psychoactive substances was observed,
(5, metab), mazindol (10), harmine (10),
with 79.9 % of OF samples containing two or more psychoactive sub­
harmaline (10), PCP (0.1), morphine (1), 6-
MAM (1, metab), heroin (1), codein (1), stances; 34.2 % had two, 23.4 % had three and others contained up to
oxycodone (10), hydrocodone (10), seven psychoactive substances (Fig. 2). The chi-square test for
meperidine (0.1), methadone (10), comparing the number of psychoactive substances detected per sample
amitriptyline (10), bupropion (10), in parties and electronic music festivals showed a significant difference
hydroxybupropion (10, metab), citalopram
(10), desmethylcitalopram (10, metab),
desipramine (10), doxepin (10), fluoxetine
(10), imipramine (10), mirtazapine (10),
nortriptyline (10), paroxetine (10),
sertraline (10), trazodone (10), quetiapine
(10), venlafaxine (10), O-
desmethylvenlafaxine (10, metab),
verapamil (10), carbamazepine (10),
clenbuterol (10), clomipramine (10),
haloperidol (10), 7-aminoclonazepam (10,
metab), 7-aminoflunitrazepam (10, metab),
bromazepam (10), flurazepam (10),
chlordiazepoxide (10, metab), midazolam
(10), medazepam (10), oxazepam (10),
nitrazepam (10), clonazepam (10),
flunitrazepam (10), alprazolam (10),
diazepam (10), nordiazepam (10, metab),
temazepam (10), lorazepam (10), clobazam
(10), duloxetine (10), levamisole (10), Fig. 1. Prevalence (%) of each New Psychoactive Substance detected in 181
ethylphenidate (10), psilocin (10), LSA (1), NPS-positive oral fluid samples from parties and electronic music festivals. A
sildenafil (10), lidocaine (10), benzocaine detailed scale is shown in the grey rectangle to better represent the results
(10) between 25C-NBOH and mCPP analytes.

3
K.F. da Cunha et al.
Fig. 2. Polydrug consumption (%) of illicit psychoactive substances observed in
(A) 462 oral fluid samples and (B) divided by event type (231 samples collected
in parties and 231 collected in electronic music festivals).
between the groups (x2(7, N = 462) = 77.79, p < 0.0001). At festivals,
the highest prevalences were for three (27.3 %) and four (24.7 %)
substances per sample, whereas at parties primarily two (44.2 %) or one
substance (27.3 %) was found. Drug prevalence at each type of event is
also detailed in Fig. 2. Of 16 cocaine-positive samples (6.9 %) from
parties, 56.3 % also included the antiparasitic levamisole, a common
cocaine adulterant, and 12.5 % contained ketamine. At electronic music
festivals, 69.3 % of 62 cocaine-positive samples (26.8 %) were also
levamisole-positive, and 75.8 % contained NPS (89.4 % ketamine, 10.6
% N-ethylpentylone, 6.4 % methylone, 6.4 % eutylone, 6.4 % dipenty­
lone, 2.1 % MDPV or 2.1 % 25I-NBOH). Of 9 methamphetamine-positive
samples (1.9 %), 44.4 % also had an NPS (50.0 % ketamine, 50.0 %
methylone and 25.0 % eutylone). Antidepressant drugs and/or benzo­
diazepines were detected in 7.4 % of samples, concurrent with other
psychoactive substances.
When comparing the illicit psychoactive substance endorsed by the
participant in the questionnaire and the OF toxicological result, there
was agreement in 143 (31.0 %) of the paired 462 samples. In 291
samples (63.0 %), at least one identified substance did not correspond to
the participant self-report. In these discordant results, 154 (52.9 %)
contained a NPS not mentioned in the questionnaire. Reported sub­
stances not analytically confirmed included LSD (69), THC (17), psilocin
(8), DMT (6), mescaline (6), cocaine (4), MDMA (2), Salvia divinorum
(1), and methamphetamine (1). In 28 samples, there was total
disagreement between self-report and toxicological results, with 28.6 %
of these containing a NPS (Fig. 3). There was some disagreement be­
tween results in 56.3 % of party samples and 81.9 % of samples from
festivals.
Only 5 % of volunteers reported NPS consumption, including two
who acknowledged consumption of two different NPS. Ketamine was the
most reported NPS (n = 23), predominantly by festival participants
(91.3 %). Although consumption of 2C-B gel caps was reported by a
volunteer, LC–MS/MS identified 25I-NBOH and 5-MeO-MiPT. One
NBOH ingestion that had occurred almost 24 h before sample collection
was not confirmed by laboratory analysis.
Of 11 methylphenidate-positive samples, two volunteers reported
use only for recreational purposes rather than treatment for attention
4
Drug and Alcohol Dependence 227 (2021) 108962
Fig. 3. ‘Agreement’, ‘some agreement’, and ‘disagreement’ obtained between
reported substances vs. oral fluid LC–MS/MS analysis. Oral fluid samples con­
taining New Psychoactive Substances are in light grey.
deficit hyperactivity disorder. Complete data on all detected and re­
ported drugs from NPS-positive samples are available in Table 2. On
average, 63.1 % of volunteers accessed their respective toxicological
results within two months of collection.
4. Discussion
In Australia in 2013, amphetamine, cannabis and polydrug users
reporting licit drug use for non-medical purposes and/or illicit drug
consumption in the last year, used synthetic cannabinoids and other NPS
more frequently than other drug users (Sutherland et al., 2018). Poly­
drug users were significantly younger and prone to more risky behaviors
under the influence of drugs and/or alcohol (e.g., driving a motor
vehicle). NPS use in the last six months was evaluated in 4122 regular
psychostimulants users between 2010–2015 (Sutherland et al., 2016).
Of these, 41.9 % reported NPS consumption, with prevalence varying
over time: 18.5 % used synthetic cathinones in 2010 decreasing to 7.7 %
in 2015. Phenylethylamines use varied from 8 to 18.6 % and trypt­
amines use from 8 to 10.9 %. Palamar and Keyes interviewed 3582
electronic music party attendees in New York (Palamar and Keyes,
2020). In 2016 and 2019, 5.9 % and 15.3 % reported using ketamine in
the last year, respectively. LSD consumption also increased from 9.9 to
16.6 %.
Calle et al. (2019) collected blood samples from 121 intoxication
cases at an electronic music event medical station in Belgium and
identified alcohol and MDMA samples as the primary psychoactive
drugs consumed by 83 % and 54 % of drug users, respectively. Ketamine,
positive in 29.4 % our samples, was identified in 6 % and PMMA in 3 %.
Other NPS (4-fluoroamphetamine, ethylone, and α-PVP) were found in
five of 20 samples analyzed by liquid chromatography coupled to a
diode-array detector and gas chromatography coupled to mass spec­
trometry, which could have hindered NPS identification due to
decreased sensitivity achieved by these analytical techniques.
The ingestion of multiple NPS is problematic. Palamar et al. (2017)
showed an increase from 3.6 % (2007–2008) to 5.4 % (2013–2014) of
12− 34 year old ecstasy users who consumed 5–8 psychoactive sub­
stances last year. In Sweden, between 1998–2007, more than one psy­
choactive substance was identified in 65 % of accidental deaths, 79 % of
undetermined deaths, and 89 % of suicides, with ethanol as the most
prevalent substance in 6894 deaths from drugs of abuse and pharma­
ceutical intoxications (Jones et al., 2011). Consumption of two sub­
stances with similar effects on the central nervous system is riskier than
using a single substance (Assi et al., 2017). Assi et al. reviewed 20
studies of NPS effects and toxicities in above 15-year-olds and found that
in 14, NPS were combined with other substances such as alcohol, energy
drinks, tobacco, and/or conventional drugs of abuse producing adverse
effects and, in three, fatal intoxications occurred. Of 670 NPS toxicology
cases reported to the UNODC between January 2019 and April 2020, 90
% identified polydrug consumption in driving under the influence of
drugs (DUID) and 81 % of postmortem cases, with 13 and 19 % of cases
identifying more than one NPS, respectively (UNODC, 2020b).
K.F. da Cunha et al. Drug and Alcohol Dependence 227 (2021) 108962

Table 2 Table 2 (continued )

Authentic oral fluid samples containing at least one New Psychoactive Substance Sample NPS Other substances Reported drug
(NPS) by LC–MS/MS, and their matched questionnaire answers, separated by consumption
event type of parties or electronic music festivals.
41 Ketamine MDMA, MDA, THC LSD, ecstasy
Sample NPS Other substances Reported drug 42 Ketamine MDMA, THC, cocaine, Cocaine
consumption levamisole
Parties 43 Ketamine THC MDMA
1 Methylone MDMA, MDA, MDEA, MD, marijuana 44 Ketamine MDMA, MDA THC
THC, 45 25I-NBOH, 25B- THC LSD
methylphenidate, NBOH, 25C-NBOH
venlafaxine 46 Ketamine MDMA, MDA, MDEA, Ecstasy, MDMA, THC
2 Methylone MDMA, MDA, MDEA, MD THC
methamphetamine 47 Ketamine MDMA, MDA, THC MDMA, THC,
3 Ketamine, 25I- MDMA, THC, Marijuana chloroform
NBOH, 25C-NBOH venlafaxine 48 Ketamine MDMA, THC THC, chloroform
4 Ketamine, MDMA, MDA, MDEA, MD 49 Ketamine MDMA, MDA, MDEA, LSD
methylone, N- THC, methylphenidate THC, LSD
ethylpentylone 50 Ketamine MDMA, MDA, MDEA, MDMA, LSD, THC
5 N-ethylpentylone MDMA, MDA, THC MD, hashish THC, LSD
6 N-ethylpentylone MDMA, MDA LSD, ecstasy 51 Dipentylone MDMA, MDA, THC Ecstasy
7 Methylone MDMA, MDA, MDEA MD, marijuana 52 Methylone MDMA, MDA, MDEA, MDMA, LSD, THC
8 Methylone MDMA, MDA, MDEA MD THC, LSD
9 Methylone MDMA, MDA, MDEA, MD, marijuana 53 Ketamine MDMA, MDA, MDEA, MDMA, THC
THC THC
10 Mephedrone MDMA, MDA, MDEA, MD, ecstasy 54 25I-NBOH MDMA, MDA, THC, MD, LSD, marijuana
THC, amphetamine LSD, clomipramine
11 Mephedrone MDMA, MDA, THC MD, ecstasy 55 Methylone MDMA, MDA MD
12 N-ethylpentylone MDMA, MDA, MDEA, Ecstasy, marijuana
THC Electronic Music Festivals
13 Methylone MDMA, MDA, MDEA, MD, marijuana 1 Ketamine MDMA, MDA, MDEA, LSD, cocaine, ecstasy,
THC THC, LSD, cocaine, marijuana
14 N-ethylpentylone MDMA, MDEA, THC, MD, marijuana, LSD, levamisole
LSD, cocaine, cocaine 2 Ketamine MDMA, THC, LSD Cocaine
levamisole, 3 N-ethylpentylone, MDMA, MDA, THC, LSD, MD
mirtazapine dipentylone LSD
15 Ketamine MDMA, MDA, THC MD 4 Ketamine, N- MDMA, MDA, LSD LSD, MD, marijuana
16 Methylone MDMA, MDA, MDEA MDMA, ecstasy ethylpentylone
17 Ketamine MDMA, MDA, MDEA Ecstasy 5 Ketamine MDMA, MDA, THC, LSD, MD, ecstasy,
18 Ketamine MDMA, MDA, MDEA Ecstasy LSD, cocaine, cocaine, mescaline
19 Methylone MDMA, MDA, MDEA, Marijuana, ecstasy, levamisole
THC MD 6 Ketamine MDMA, MDA, THC Ecstasy
20 Methylone THC, LSD LSD, marijuana 7 N-ethylpentylone MDMA, MDA, THC LSD, MD, ecstasy
21 Ketamine MDMA, MDEA, THC, Marijuana, MD, LSD 8 Ketamine, N- MDMA, THC, cocaine, LSD, cocaine
LSD ethylpentylone levamisole
22 2C-X LSD 9 Dipentylone MDMA, THC LSD
23 Ketamine MDMA, MDA, MDEA, LSD 10 N-ethylpentylone MDMA LSD, MD, ecstasy
THC, LSD 11 N-ethylpentylone MDMA, MDA, THC LSD, MD, ecstasy
24 25C-NBOH MDMA, THC, Hashish 12 Ketamine, N- MDMA, MDA, MDEA, LSD, MD, ecstasy,
methylphenidate ethylpentylone, THC, LSD, cocaine marijuana
25 Ketamine MDMA, MDA, MDEA, MD dipentylone
THC, cocaine, 13 Ketamine MDMA, THC, LSD LSD, MD, marijuana
levamisole 14 Ketamine MDMA, THC, LSD LSD, MD, marijuana
26 Ketamine MDMA, MDA, MDEA, Ecstasy, MD 15 Methylone, 25C- MDMA, MDA, MDEA, LSD, MD, marijuana
THC NBOH THC
27 Ketamine MDMA, MDA, MDEA, LSD, marijuana 16 Ketamine MDMA, THC, cocaine, Cocaine
THC levamisole
28 Ketamine MDMA, MDA, MDEA, Ecstasy 17 Ketamine MDMA, MDA, MDEA, MDMA, mushroom
THC THC, cocaine,
29 Ketamine MDMA, MDA, MDEA, Ecstasy levamisole, ephedrine
THC 18 Ketamine MDMA, THC, psilocin LSD, marijuana,
30 Ketamine MDMA, MDA, THC MDMA, THC mushroom
31 Ketamine, N- MDMA, MDA, MDEA, Ecstasy, ketamine, 19 Ketamine, mCPP MDMA, MDA, THC LSD, MDMA
ethylpentylone, THC, LSD THC, LSD 20 Ketamine, MDMA, MDA, THC, LSD, marijuana
dipentylone dipentylone LSD
32 Ketamine MDMA, THC MDMA, THC 21 Ketamine MDMA, MDA, THC, MD
33 Ketamine MDMA, MDA, THC MDMA cocaine
34 Ketamine MDMA, MDEA, THC, MDMA, ketamine 22 Ketamine MDMA, MDA, MDEA, Ketamine, ecstasy,
LSD THC LSD, MDMA,
35 Ketamine MDMA, MDA, THC MDMA poppers, marijuana
36 Ketamine MDMA, MDA, LSD LSD 23 Ketamine MDMA, MDA, MDEA, LSD, MDMA,
37 N-ethylpentylone MDMA, MDA, THC, Ecstasy, hashish THC, LSD, citalopram, marijuana/hashish
LSD alprazolam
38 N-ethylpentylone, MDMA, THC, LSD LSD 24 Ketamine MDMA, MDA, THC, Ketamine, ecstasy,
dipentylone cocaine, levamisole LSD
39 Ketamine MDMA, MDA, MDEA, Ecstasy, LSD 25 Ketamine MDMA, MDA, THC, LSD, ecstasy
LSD LSD
40 25I-NBOMe LSD LSD 26 N-ethylpentylone MDMA, THC
(continued on next page)

5
K.F. da Cunha et al. Drug and Alcohol Dependence 227 (2021) 108962

Table 2 (continued ) Table 2 (continued )

Sample NPS Other substances Reported drug Sample NPS Other substances Reported drug
consumption consumption

LSD, marijuana/ 63 Ketamine MDMA, MDA, THC, Cocaine, marijuana

hashish cocaine, levamisole
27 Ketamine MDMA, MDA, THC LSD, MD, ecstasy, 64 Ketamine MDMA, MDA, cocaine, LSD, cocaine, MD
marijuana/hashish levamisole
28 Dipentylone MDMA, MDA, THC, LSD, MD, marijuana 65 Ketamine MDMA, MDA, THC, Cocaine, marijuana
cocaine, levamisole cocaine
29 Ketamine MDMA, MDA, THC, LSD, marijuana, 66 Ketamine MDMA, THC, LSD LSD, MD, marijuana
cocaine MDMA, cocaine 67 Ketamine MDMA, MDA, THC, Cocaine, marijuana,
30 Methylone MDMA, MDA, THC, LSD, MD, ecstasy, cocaine, levamisole mushroom
LSD, psilocin marijuana, 68 Ketamine MDMA, MDA, LSD, MD
mushroom citalopram
31 Ketamine MDMA, MDA, THC Ketamine, MD, 69 Ketamine MDMA, MDA, THC, MD, LSD, marijuana
marijuana citalopram
32 Ketamine MDMA, MDA, MDEA, MD, hashish 70 Ketamine, MDMA, MDA, MDEA, MD
THC methylone THC, LSD, cocaine,
33 Ketamine MDMA, MDA, MDEA, MD, marijuana/ levamisole,
THC, LSD hashish methamphetamine
34 Ketamine, N- MDMA, MDA, THC, Mescaline, THC 71 Methylone MDMA, MDA, MDEA, MD, hashish
ethylpentylone, cocaine, levamisole THC, quetiapine
dipentylone 72 Ketamine MDMA, MDA, THC, Cocaine, MD,
35 Ketamine MDMA, THC, LSD, LSD, mescaline, DMT, cocaine, levamisole ketamine, marijuana
cocaine, levamisole cocaine, ketamine, 73 Ketamine, 4- MDMA, THC MD, marijuana
THC chloroethcathinone
36 Ketamine MDMA, MDA, THC, LSD, MD, THC 74 Ketamine MDMA, MDA, MDEA, MD
LSD THC
37 Ketamine MDMA, MDA, THC LSD, MD, THC 75 Ketamine MDMA, MDA, THC, LSD, MD, marijuana,
38 Ketamine MDMA, THC, harmine Changa, LSD, MD, LSD changa
marijuana 76 Ketamine MDMA, THC, LSD LSD, marijuana
39 Ketamine MDMA, MDA, THC, Changa, LSD, MD, 77 Ketamine, MDMA, MDA, MDEA, MDMA, ecstasy,
harmine marijuana methylone THC marijuana
40 Ketamine MDMA, MDA, MDEA, Changa, marijuana/ 78 Ketamine MDMA, MDA, MDEA, MD, ecstasy,
THC, harmine hashish, MD THC marijuana
41 Ketamine MDMA, MDA, THC, Changa, LSD, MD, 79 Ketamine MDMA, MDA MD
harmine marijuana 80 25I-NBOH Ecstasy, marijuana
42 Ketamine, MDPV MDMA, MDA, THC, cocaine, ketamine, 81 Ketamine MDMA, MDA, MDEA, Ecstasy, MD,
LSD, cocaine, mescaline, LSD, THC cocaine ketamine, cocaine
levamisole 82 Ketamine, MDMA, MDA, MDEA, MD, ecstasy,
43 Ketamine, N- MDMA, MDA, MDEA, MDMA, LSD, ecstasy methylone THC marijuana
ethylpentylone THC 83 Ketamine MDMA, MDA, MD, ecstasy,
44 Ketamine MDMA, MDA, THC, MDMA, cocaine citalopram mushroom,
cocaine, levamisole mescaline, marijuana
45 Ketamine MDMA, MDA, THC LSD, MDMA, hashish 84 Ketamine MDMA, MDA, MDEA, LSD, ecstasy,
46 Ketamine MDMA, THC, LSD LSD THC, LSD marijuana
47 Ketamine MDMA, MDA, MDEA, MDMA 85 Ketamine MDMA, MDA, MDEA, MDMA, LSD, cocaine,
THC, citalopram THC, LSD, cocaine, marijuana, ketamine,
48 Ketamine MDMA, MDA MDMA diazepam mushroom
49 Ketamine MDMA, MDA, psilocin Ecstasy, mushroom, 86 Ketamine MDMA, MDA, MDEA, LSD, ecstasy, cocaine,
LSD THC, cocaine, ketamine, marijuana
50 Ketamine MDMAMDA, THC, MD, marijuana levamisole
cocaine 87 Ketamine, N- MDMA, MDA, MD, ecstasy
51 Ketamine MDMA, MDA, THC, Cocaine, ecstasy, ethylpentylone citalopram
cocaine, levamisole marijuana 88 Ketamine MDMA, MDA, THC, Cocaine, LSD,
52 Ketamine MDMA, MDA, THC, Cocaine, MD, LSD, cocaine, ecstasy, MD,
cocaine, levamisole marijuana levamisole marijuana
53 Ketamine MDMA, MDA, THC, Mescaline, LSD, 89 Ketamine, 25I- MDMA, MDA, MDEA, MD, ecstasy
LSD marijuana/hashish NBOH THC
54 Ketamine MDMA, THC, Marijuana, 90 Ketamine Changa
methamphetamine, methamphetamine 91 Ketamine MDMA, THC Ecstasy, MD,
amphetamine ketamine, THC
55 Ketamine MDMA, MDA, THC Cocaine, ketamine, 92 Ketamine MDMA, MDA Ketamine, ecstasy,
cocaine MD, LSD cocaine, marijuana
56 Ketamine MDMA, MDA, THC, Cocaine, ketamine, 93 Ketamine MDMA, MDA, THC MD, marijuana
cocaine MD, marijuana 94 Eutylone MDMA, MDA, THC Ecstasy
57 Ketamine MDMA, MDA, THC MD, ketamine, 95 Ketamine MDMA, MDA, THC, Ecstasy, marijuana
marijuana LSD
58 Ketamine, MDMA, MDA, THC LSD, MDMA, 96 Ketamine MDMA, THC, LSD, Cocaine, LSD,
methylone marijuana cocaine, levamisole marijuana
59 Ketamine MDMA, MDA, THC, Cocaine, MD, 97 Ketamine MDMA, MDA, MDEA, MD, ecstasy,
cocaine, levamisole marijuana THC marijuana
60 Methylone MDMA, MDA, MDEA, LSD, MD, mushroom, 98 Methylone MDMA, MDA MDEA, MD, ecstasy, LSD,
THC, psilocin cocaine, marijuana THC cocaine, marijuana
61 Ketamine MDMA, MDA, THC, MD, marijuana, 99 Ketamine MDMA, MDA, THC, MD, ecstasy, LSD,
cocaine, levamisole cocaine LSD, cocaine, cocaine, hashish,
62 Ketamine MDMA, THC, cocaine, Cocaine, MD, levamisole ethyl chloride
levamisole marijuana, poppers 100 Ketamine
(continued on next page)

6
K.F. da Cunha et al. Drug and Alcohol Dependence 227 (2021) 108962

Table 2 (continued ) environment. Ketamine was present in 13 % of samples collected at

Sample NPS Other substances Reported drug parties, and in almost 46 % at festivals, with overall more NPS con­
consumption sumption at festivals.
MDMA, MDA, MDEA, MD, LSD, cocaine,
According to the Brazilian Federal Police, reports issued for synthetic
THC, LSD marijuana drugs increased from 7 to 19 % from 2017 to 2018, with MDMA, N-
101 Ketamine MDMA, MDA, THC, MD, marijuana, ethylpentylone, MDA (3,4-methylenedioxyamphetamine), LSD, and 25I-
cocaine, levamisole cocaine NBOH being the five most reported substances. Although reports of
102 Ketamine MDMA, MDA, THC, LSD, MD, cocaine,
conventional synthetic drugs were the majority (almost 70 % in 2018),
LSD, cocaine, marijuana
levamisole there was still an increase of 112 % in reports on synthetic cathinones
103 Methylone, MDMA, MDA, THC, Ecstasy, cocaine, (Brazilian Federal Police, 2020).
eutylone, 25I-NBOH cocaine, codeine marijuana MDMA was the main active ingredient in ecstasy pills (Palamar et al.,
104 Ketamine MDMA, MDA, LSD LSD, MD, marijuana 2017). In Brazil, ecstasy consumption paralleled increased ecstasy pill
105 Ketamine, 25I- MDMA 2C-G, salvia,
NBOH, 5-MeO-MiPT ketamine, MD,
seizures. In the Southern Brazilian Santa Catarina state, 3472 ecstasy
marijuana seizures occurred from 2011 to 2017; 87 in 2011, and 973 in 2017
106 Methylone, 5-MeO- MDMA, MDA, MDEA, LSD, MDMA, (Souza et al., 2020). Up to six substances (caffeine, clobenzorex, keta­
MiPT THC, LSD marijuana mine, mCPP, amphetamine, 2C-B, procaine, MDEA (N-ethyl-3,4-meth­
107 Methylone MDMA, MDA, MDEA, MD, LSD, ecstasy,
ylenedioxyamphetamine), lidocaine, ethylone, and N-ethylpentylone)
THC, LSD, cocaine, cocaine, marijuana
levamisole were identified in 30 % of ecstasy seizures. N-ethylpentylone was found
108 Ketamine MDMA, MDA, THC, LSD, MD, marijuana in 3.2 % and 13 % of confiscated pills as a substitute for MDMA in 2016
LSD and 2017, respectively. mCPP is also a metabolite of trazodone; how­
109 Ketamine, 5-MeO- THC, LSD LSD, marijuana ever, trazodone was monitored in the method and was not detected in
MiPT
110 Ketamine MDMA, MDA, THC, DMT, LSD, MD
the same sample as mCPP. In addition, other publications reported
LSD mCPP in combination with MDMA in ecstasy pills, including seized
111 Eutylone MDMA, MDA, MDEA, MD, ecstasy tablets in Brazil (Bossong et al., 2010; Souza et al., 2020). Thus, mCPP
cocaine, levamisole, positive samples are more likely due to recreational abuse than trazo­
methamphetamine,
done prescription.
amphetamine
112 Methylone MDMA, MDA, MDEA, LSD, MDMA, changa, LSD was the sixth most prevalent psychoactive substance among the
THC, LSD, cocaine, cocaine, marijuana samples analyzed in our study (23.6 %), generally present on blotter
levamisole papers. NBOMes, NBOHs, furanylfentanyl, and the synthetic cannabi­
113 Methylone MDMA, MDA, THC Ecstasy, MD, noid AMB-FUBINACA were also identified in confiscated blotter papers
marijuana
in Brazil (Barbosa et al., 2019; de Souza Boff et al., 2020; Rodrigues de
114 Ketamine MDMA, MDA, cocaine, Cocaine, MD
levamisole Morais et al., 2020). Among volunteers participating in this research,
115 Ketamine MDMA, MDA, MDEA, Cocaine, ketamine, one reported LSD consumption, although toxicological analysis of his OF
THC, LSD, cocaine ecstasy, marijuana, was positive for 25I-NBOH, 25C-NBOH and 25B-NBOH, not LSD.
LSD
The choice of OF as an alternative matrix to blood and urine made
116 Ketamine MDMA, THC Ecstasy, marijuana
117 Ketamine, MDMA, MDA, MDEA, MD, LSD, ketamine, onsite collection much more feasible and identified recent psychoactive
methylone THC, LSD marijuana substance use. Emotional state, level of hydration, and/or consumption
118 Ketamine MDMA, MDA, MDEA, LSD, MD, ecstasy, of stimulants, including MDMA, amphetamines and cannabis, can
LSD ketamine reduce salivation, prolonging OF collection time (Drummer, 2006).
119 Ketamine MDMA, THC, cocaine, NBOH, MD,
Prolonged collection was primarily observed in sample collections at
bupropion ketamine, cocaine,
marijuana festivals, where drugs are consumed over many hours, where room and
120 Ketamine MDMA, MDA, THC ketamine, ecstasy, body temperatures may be elevated, and where access to water is often
MD, LSD, hashish restricted.
121 Ketamine MDMA, THC, cocaine, Cocaine, MD,
Biological matrix analysis cannot distinguish whether multiple drugs
levamisole marijuana
122 Ketamine MDMA, MDA, MDEA, MD, LSD,
in the same sample were due to the consumption of one substance with
THC, cocaine methamphetamine, several adulterants or polydrug use of several substances over a short
hashish period of time. One volunteer reported “MD” consumption (crystal
123 Ketamine MDMA, MDA, MDEA, MD, ecstasy, LSD, MDMA), whereas toxicological analysis identified not only MDMA, but
THC, LSD marijuana
also N-ethylpentylone, ketamine, and methylone. Two volunteers re­
methylphenidate,
bupropion ported MDMA consumption 30− 60 min before OF collection, but only
124 Ketamine, MDMA, MDA, MDEA, LSD, changa, MD, MDA, a metabolite of MDMA and a drug with of its own psychoactive
methylone, eutylone THC, LSD, cocaine, cocaine, marijuana properties was observed (Monks et al., 2004).
levamisole, harmine
One study limitation was utilization of a target analysis method,
125 Ketamine MDMA, MDA, THC, LSD, psilocin,
LSD venlafaxine marijuana
based on a triple quadrupole mass spectrometer, operating in multiple
126 Ketamine MDMA, MDA, THC, Ketamine, ecstasy, reaction monitoring (MRM) mode, allowing identification of substances
cocaine, levamisole cocaine, marijuana previously optimized with certified standards, rather than a non-
MD: MDMA consumed as crystal/powder form; Ecstasy: MDMA consumed as pill targeted high resolution mass accuracy method. Although our method
form; Changa: smoking mixture containing DMT and β-carbolines. contains more than 100 NPS and conventional drugs of abuse, some
substances may not have been identified within the scope of the method
There are few data on NPS consumption in Brazil, generally limited library. This method is constantly updated to minimize false negative
to police seizures or intoxication cases (Costa et al., 2019). Our study results based upon Brazilian Federal Police publications and, mainly,
was the first to assess the consumption of these new and important drugs NPS warning systems (Center for Forensic Science Research and Edu­
of abuse, including data where NPS are most frequently consumed. The cation, 2021; EMCDDA, 2021a; National Forensic Laboratory Informa­
types of psychoactive substances identified at parties and electronic tion System (NFLIS), 2021; UNODC, 2021). Another volunteer reported
music festivals were similar; however, the prevalence differed by NBOH consumption almost 24 h prior to collection that was not
analytically confirmed, likely due to the elapsed time and/or low dose.

7
K.F. da Cunha et al.
The NBOH consumed may not be among the four analogs included in the
analytical method; Brazilian Federal Police reported five different NBOH
from seizures in 2018 (Brazilian Federal Police, 2020).
In our last collection during the 2nd semester 2019, eutylone, a
synthetic cathinone, was identified for the first time in four (3.6 %) of
112 samples (Supplementary Table 3). Eutylone was present in over 50
forensic toxicological cases (postmortem and DUID) between September
2019 and March 2020 in the USA (CFSRE, 2020), reinforcing the
importance of continuously updating the analytical method to include
NPS identified in other countries.
Due to varied NPS potency, it is difficult to estimate the instrumental
sensitivity required for new NPS identification, especially in OF samples
as few NPS publications report OF data. Even for conventional drugs of
abuse, different detection and quantification cutoffs are recommended:
the Substance Abuse and Mental Health Services Administration
(SAMHSA) sets 50 ng/mL as the cutoff for MDMA confirmatory tests in
OF, whereas Driving Under the Influence of Drugs, Alcohol and Medi­
cines (DRUID) recommends 25 ng/mL (Bosker and Huestis, 2009).
Highly sensitive limits of detection were employed in this study to avoid
false negative results. However, the lack of data from controlled NPS
administration studies, does not allow determination of detection win­
dows for each NPS class.
Differences between questionnaire responses and toxicological ana­
lyses may be due to (a) a volunteer’s decision not to report a substance,
(b) unknown intake or (c) accuracy in remembering drug use. These
factors contribute to the differences in analytical results and self-reports
between festivals (81.8 %) and parties (56.3 %). Johnson et al. (2009)
also addressed the predisposition to report consumption depending
upon consumed substance, after observing a greater underreporting for
cocaine (67 %) and amphetamines (52 %) compared to cannabis (32 %).
Only 12.7 % of sample donors with NPS-positive samples, reported NPS
consumption. Among 60 volunteers at electronic music festivals who
reported using MDMA in the past week, 47 % had NPS in their biological
samples, whereas 15 % were MDMA-positive and 17 %, MDMA- and
NPS-positive (Mohr et al., 2018). In the present study, of 39.2 %
NPS-positive samples, only 4.4 % did not contain MDMA.
High rates of conventional synthetic drug adulteration/substitution,
mainly with NPS, demonstrates the vulnerability and increased risk of
consumers for severe intoxications (Costa et al., 2019; Krotulski et al.,
2018; Suzuki et al., 2015; Swanson et al., 2017; Wilde et al., 2020).
Other risks from psychoactive substance abuse include aggressive
behavior, injuries, unprotected sex, and DUID (EMCDDA, 2016). Of 218
NPS identified in 181 samples in this study, 70.6 % were in samples from
electronic music festivals, which may indicate an increased risk of
consumption in this context.
Knowing psychoactive substances consumed in a country can assist
in formulating more effective public health policies and/or adopt stra­
tegies to reduce risks and harms associated with their consumption.
Access to toxicological analyses by most volunteers shows there is in­
terest in knowing substances consumed.
5. Conclusion
We present the first data on NPS consumption in Brazil based on
onsite OF collection at parties and electronic music festivals. Ketamine
(29.4 %), methylone (6.1 %), and N-ethylpentylone (4.1 %) were the
most prevalent NPS in 462 samples collected between September 2018
and January 2020. Although 39.2 % of OF samples were positive for
NPS, only 5 % of 462 volunteers reported NPS consumption. Users
inadvertently consume NPS suggesting that risk and harm reduction
strategies are needed to prevent intoxication cases.
Ethical approval
All procedures performed in studies involving human participants
were in accordance with the ethical standards of the University of
8
Drug and Alcohol Dependence 227 (2021) 108962
Campinas committee (Comitê de Ética em Pesquisa da UNICAMP—CEP;
CAAE 88770318.0.0000.5404) and with the 1964 Helsinki Declaration.
Contributors
All authors participated in the review of this report research. Kelly
Francisco da Cunha performed the sample collections, formal analysis,
visualization, and wrote the original draft. Marilia Santoro Cardoso,
Ana Carolina Furiozo Arantes, Pedro Henrique Piras Coser, Lucas
de Noronha Lima and Maria Angélica De Castro Comis performed
the sample collections. Karina Diniz Oliveira conducted the resources
and conceptualization. Marilyn A Huestis critically reviewed and
revised the manuscript. Jose Luiz Costa conducted the conceptualiza­
tion, resources and methodology of this project, in addition to writing
and revising the original and final manuscript.
Declaration of Competing Interest
The authors report no declarations of interest.
Acknowledgements and funding source
The authors thank the São Paulo Research Foundation – FAPESP
(process Nos.2017/02147-0,2018/00432-1, and2018/11849-0), the­
Coordination for the Improvement of Higher Education Personnel –
CAPES (Finance Code001) and theFundo de Apoio ao Ensino, Pesquisa e
Extensão da UNICAMP‒FAEPEX (process No.14379-18) by the financial
support; the Volunteer Research Service of the National Secretariat on
Drug Policy (SVP-SENAD, Brazil) by support; and the Espaço da Escrita –
Pró-Reitoria de Pesquisa – UNICAMP – for the language services
provided.
Appendix A. Supplementary data
Supplementary material related to this article can be found, in the
online version, at doi:https://doi.org/10.1016/j.drugalcdep.2021.10
8962.
References
Adams, A.J., Banister, S.D., Irizarry, L., Trecki, J., Schwartz, M., Gerona, R., 2017.
“Zombie” outbreak caused by the synthetic cannabinoid AMB-FUBINACA in New
York. N. Engl. J. Med. 376, 235–242. https://doi.org/10.1056/NEJMoa1610300.
Assi, S., Gulyamova, N., Ibrahim, K., Kneller, P., Osselton, D., 2017. Profile, effects, and
toxicity of novel psychoactive substances: a systematic review of quantitative
studies. Hum. Psychopharmacol. Clin. Exp. 32, e2607. https://doi.org/10.1002/
hup.2607.
Barbosa, L., de, M., Santos, J.M., de Morais, D.R., Nimtz, A.V., Eberlin, M.N., de
Oliveira, M.F., Costa, J.L., 2019. Fast UHPLC-MS/MS method for analysis of
furanylfentanyl in different seized blotter papers. Drug Test. Anal. 11, 178–183.
https://doi.org/10.1002/dta.2472.
Bosker, W.M., Huestis, M.A., 2009. Oral fluid testing for drugs of abuse. Clin. Chem. 55,
1910–1931. https://doi.org/10.1373/clinchem.2008.108670.
Bossong, M.G., Brunt, T.M., Van Dijk, J.P., Rigter, S.M., Hoek, J., Goldschmidt, H.M.J.,
Niesink, R.J.M., 2010. mCPP: an undesired addition to the ecstasy market.
J. Psychopharmacol. 24, 1395–1401. https://doi.org/10.1177/0269881109102541.
Brazilian Federal Police, 2020. 2018 Report - Synthetic Drugs. https://www.gov.br/pf/pt
-br/acesso-a-informacao/acoes-e-programas/relatorio-de-drogas-sinteticas-2018.
Calle, P., Maudens, K., Lemoyne, S., Geerts, S., Van Sassenbroeck, D., Jensen, P., Van
Overloop, J., Deconinck, E., Blanckaert, P., 2019. Lessons to be learned from
toxicological analyses in intoxicated patients and seized materials at an electronic
music dance festival. Forensic Sci. Int. 299, 174–179. https://doi.org/10.1016/j.
forsciint.2019.03.047.
Center for Forensic Science Research and Education, 2020. Eutylone (bk-EBDB) and
Benzylone (BMDP): Increasing Prevalence of New Synthetic Stimulants in the United
States. https://www.npsdiscovery.org/wp-content/uploads/2020/03/Public-Alert
_Eutylone_Benzylone_NPS-Discovery_033120.pdf.
Center for Forensic Science Research and Education. NPS discovery - Trend reports.
https://www.npsdiscovery.org/.
Costa, J.L., Cunha, K.F., Lanaro, R., Cunha, R.L., Walther, D., Baumann, M.H., 2019.
Analytical quantification, intoxication case series, and pharmacological mechanism
of action for N-ethylnorpentylone (N-ethylpentylone or ephylone). Drug Test. Anal.
11, 461–471. https://doi.org/10.1002/dta.2502.
K.F. da Cunha et al.
da Cunha, K.F., Oliveira, K.D., Huestis, M.A., Costa, J.L., 2020. Screening of 104 new
psychoactive substances (NPS) and other drugs of abuse in oral fluid by LC–MS-MS.
J. Anal. Toxicol. 44, 697–707. https://doi.org/10.1093/jat/bkaa089.
de Souza Boff, B., Silveira Filho, J., Nonemacher, K., Driessen Schroeder, S., Dutra
Arbo, M., Rezin, K.Z., 2020. New psychoactive substances (NPS) prevalence over
LSD in blotter seized in State of Santa Catarina, Brazil: a six-year retrospective study.
Forensic Sci. Int. 306, 110002 https://doi.org/10.1016/j.forsciint.2019.110002.
DEA, 2019. National Drug Threat Assessment. https://www.dea.gov/sites/default/fil
es/2020-01/2019-NDTA-final-01-14-2020_Low_Web-DIR-007-20_2019.pdf.
Desrosiers, N.A., Huestis, M.A., 2019. Oral fluid drug testing: analytical approaches,
issues & interpretation of results. J. Anal. Toxicol. 43, 415–443. https://doi.org/
10.1093/jat/bkz048.
Drummer, O.H., 2006. Drug testing in oral fluid. Clin. Biochem. Rev. 27, 147–159.
EMCDDA, 2016. Health Responses to New Psychoactive Substances. https://www.emcdd
a.europa.eu/system/files/publications/2812/TD0216555ENN.pdf.
EMCDDA, 2019. European Drug Report - Trends and Developments. http://www.
emcdda.europa.eu/system/files/publications/11364/20191724_TDAT19001ENN
_PDF.pdf.
EMCDDA. EU Early Warning System. http://www.emcdda.europa.eu/publications/topi
c-overviews/eu-early-warning-system.
EMCDDA. New Psychoactive Substances (NPS). https://www.emcdda.europa.eu/topi
cs/nps_en.
Fernández-Calderón, F., Cleland, C.M., Palamar, J.J., 2018. Polysubstance use profiles
among electronic dance music party attendees in New York City and their relation to
use of new psychoactive substances. Addict. Behav. 78, 85–93. https://doi.org/
10.1016/j.addbeh.2017.11.004.
Hedegaard, H., Miniño, A.M., Warner, M., 2020. Drug Overdose Deaths in the United
States, 1999–2018. NCHS Data Brief, 356. https://www.cdc.gov/nchs/products/d
atabriefs/db356.htm.
Hondebrink, L., Nugteren-van Lonkhuyzen, J.J., Hunault, C.C., Berg, J., Gouwe, D.,
Riel, A.J.H.P., 2020. New psychoactive substances (NPS) in the Netherlands:
occurrence in forensic drug samples, consumer drug samples and poisons center
exposures between 2013 and 2017. Addiction 115, 716–725. https://doi.org/
10.1111/add.14868.
Immunalysis, 2020. Quantisal Collection Flyer - How to Collect a Specimen with
Quantisal. https://immunalysis.com/products/oral-fluid/quantisal/collector-tra
ining/.
Johnson, M.B., Voas, R.A., Miller, B.A., Holder, H.D., 2009. Predicting drug use at
electronic music dance events: self-reports and biological measurement. Eval. Rev.
33, 211–225. https://doi.org/10.1177/0193841X09333253.
Jones, A.W., Kugelberg, F.C., Holmgren, A., Ahlner, J., 2011. Drug poisoning deaths in
Sweden show a predominance of ethanol in mono-intoxications, adverse drug-
alcohol interactions and poly-drug use. Forensic Sci. Int. 206, 43–51. https://doi.
org/10.1016/j.forsciint.2010.06.015.
Krotulski, A.J., Papsun, D.M., De Martinis, B.S., Mohr, A.L.A., Logan, B.K., 2018. N-ethyl
pentylone (ephylone) intoxications: quantitative confirmation and metabolite
identification in authentic human biological specimens. J. Anal. Toxicol. 42,
467–475. https://doi.org/10.1093/jat/bky025.
Misailidi, N., Papoutsis, I., Nikolaou, P., Dona, A., Spiliopoulou, C., Athanaselis, S., 2018.
Fentanyls continue to replace heroin in the drug arena: the cases of ocfentanil and
carfentanil. Forensic Toxicol. 36, 12–32. https://doi.org/10.1007/s11419-017-
0379-4.
Mohr, A.L.A., Friscia, M., Yeakel, J.K., Logan, B.K., 2018. Use of synthetic stimulants and
hallucinogens in a cohort of electronic dance music festival attendees. Forensic Sci.
Int. 282, 168–178. https://doi.org/10.1016/j.forsciint.2017.11.017.
Drug and Alcohol Dependence 227 (2021) 108962
Monks, T.J., Jones, D.C., Bai, F., Lau, S.S., 2004. The role of metabolism in 3,4-
(±)-methylenedioxyamphetamine and 3,4-(±)-methylenedioxymethamphetamine
(ecstasy) toxicity. Ther. Drug Monit. 26, 132–136. https://doi.org/10.1097/
00007691-200404000-00008.
National Forensic Laboratory Information System (NFLIS). Published reports htt
ps://www.nflis.deadiversion.usdoj.gov/drug.xhtml, 2021.
Palamar, J.J., Keyes, K.M., 2020. Trends in drug use among electronic dance music party
attendees in New York City, 2016–2019. Drug Alcohol Depend. 209, 107889 https://
doi.org/10.1016/j.drugalcdep.2020.107889.
Palamar, J.J., Mauro, P.M., Han, B.H., Martins, S.S., 2017. Shifting characteristics of
ecstasy users ages 12–34 in the United States, 2007–2014. Drug Alcohol Depend.
181, 20–24. https://doi.org/10.1016/j.drugalcdep.2017.09.011.
Pichini, S., Solimini, R., Berretta, P., Pacifici, R., Busardò, F.P., 2018. Acute intoxications
and fatalities from illicit fentanyl and analogues: an update. Ther. Drug Monit. 40,
38–51. https://doi.org/10.1097/FTD.0000000000000465.
Rodrigues de Morais, D., Francisco da Cunha, K., Betoni Rodrigues, T., Lanaro, R., de
Melo Barbosa, L., Jardim Zacca, J., Nogueira Eberlin, M., Costa, J.L., 2020. Triple
quadrupole–mass spectrometry protocols for the analysis of NBOMes and NBOHs in
blotter papers. Forensic Sci. Int. 309, 110184 https://doi.org/10.1016/j.
forsciint.2020.110184.
Souza Júnior, J.L., Silveira Filho, J., Boff, B.S., Nonemacher, K., Rezin, K.Z., Schroeder, S.
D., Ferrão, M.F., Danielli, L.J., 2020. Seizures of clandestinely produced tablets in
Santa Catarina, Brazil: the increase in NPS from 2011 to 2017. J. Forensic Sci. 65,
906–912. https://doi.org/10.1111/1556-4029.14237.
Sutherland, R., Peacock, A., Whittaker, E., Roxburgh, A., Lenton, S., Matthews, A.,
Butler, K., Nelson, M., Burns, L., Bruno, R., 2016. New psychoactive substance use
among regular psychostimulant users in Australia, 2010–2015. Drug Alcohol
Depend. 161, 110–118. https://doi.org/10.1016/j.drugalcdep.2016.01.024.
Sutherland, R., Peacock, A., Roxburgh, A., Barratt, M.J., Burns, L., Bruno, R., 2018.
Typology of new psychoactive substance use among the general Australian
population. Drug Alcohol Depend. 188, 126–134. https://doi.org/10.1016/j.
drugalcdep.2018.03.034.
Suzuki, J., Dekker, M.A., Valenti, E.S., Arbelo Cruz, F.A., Correa, A.M., Poklis, J.L.,
Poklis, A., 2015. Toxicities associated with NBOMe ingestion—a novel class of
potent hallucinogens: a review of the literature. Psychosomatics 56, 129–139.
https://doi.org/10.1016/J.PSYM.2014.11.002.
Swanson, D.M., Hair, L.S., Strauch Rivers, S.R., Smyth, B.C., Brogan, S.C., Ventoso, A.D.,
Vaccaro, S.L., Pearson, J.M., 2017. Fatalities involving Carfentanil and Furanyl
Fentanyl: two case reports. J. Anal. Toxicol. 41, 498–502. https://doi.org/10.1093/
jat/bkx037.
UNODC, 2020a. World Drug Report. https://wdr.unodc.org/wdr2020/.
UNODC, 2020b. Current NPS Threats, vol. III. https://www.unodc.org/documents/s
cientific/Current_NPS_Threats_Vol.3.pdf.
UNODC, 2021. Early Warning Advisory on New Psychoactive Substances. https://www.
unodc.org/LSS/Page/NPS.
Wilde, M., Sommer, M.J., Auwärter, V., Hermanns-Clausen, M., 2020. Acute severe
intoxication with cyclopropylfentanyl, a novel synthetic opioid. Toxicol. Lett. 320,
109–112. https://doi.org/10.1016/j.toxlet.2019.11.025.
Zaami, S., Giorgetti, R., Pichini, S., Pantano, F., Marinelli, E., Busardò, F.P., 2018.
Synthetic cathinones related fatalities: an update. Eur. Rev. Med. Pharmacol. Sci. 22,
268–274. https://doi.org/10.26355/eurrev_201801_14129.
Zawilska, J.B., Kacela, M., Adamowicz, P., 2020. NBOMes–highly potent and toxic
alternatives of LSD. Front. Neurosci. 14 (78), 1–15. https://doi.org/10.3389/
fnins.2020.00078.