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Neuroendocrinol Lett 2010; 31(1):101–105
ORIGINAL ARTICLE
Neuroendocrinology Letters Volume 31 No. 1 2010
Elevated urine levels of bufotenine in patients with
autistic spectrum disorders and schizophrenia
Enzo E, Roberto C, Valentina M, Natascia B,
Mara M, Marianna B, Francesco B, Pierluigi P
Department of Health Sciences, Section of Psychiatry, University of Pavia, Pavia, Italy.
Correspondence to: Enzo Emanuele, MD.
Department of Health Sciences, Section of Psychiatry,
University of Pavia, Via Bassi, 21, I-27100, Pavia, Italy.
: +39 0382 987 940; : +39 0382 526 723; -: enzo.em@libero.it
Submitted: 2009-08-06 Accepted: 2009-11-11 Published online: 2010-00-00
Key words: bufotenine; autism; schizophrenia; high-performance liquid chromatography-
mass spectrometry
Neuroendocrinol Lett 2010; 31(1):101–105 PMID: ----- NEL310110AXX © 2010 Neuroendocrinology Letters • www.nel.edu
Abstract
OBJECTIVE: Previous studies have suggested that the endogeneous psychoto-
mimetic molecule bufotenine (N-N-dimethyl-5-idroxytryptamine) may play a
role in the pathogenesis of severe mental disorders. The potential association of
bufotenine with the clinical features of autism and schizophrenia is not entirely
understood. In this study, we measured urinary levels of bufotenine in subjects
with autistic spectrum disorder (ASD), schizophrenia and healthy comparison
subjects free of psychiatric symptoms. We also sought to assess whether urine
concentrations of this molecule may be associated with the clinical characteristics
of psychiatric patients.
DESIGN: Urine bufotenine levels were measured using a high-performance liquid
chromatography-mass spectrometry (HPLC-MS) assay in young adults with
severe ASD (n=15), patients with schizophrenia (n=15), and healthy control sub-
jects (n=18). The Vineland Adaptive Behavior Scale was used to measure adaptive
behaviors in ASD individuals. The Brief Psychiatric Rating Scale (BPRS) was used
for patients with schizophrenia.
RESULTS: Urine bufotenine levels were significantly higher in ASD subjects
(3.30 ± 0.49 g/L, p<0.05) and patients with schizophrenia (4.39 ± 0.43 g/L,
p<0.001) compared with controls
(1.53 ± 0.30 g/L). Among patients with ASD, there was a significant positive
correlation between urine bufotenine and hyperactivity scores on the Vineland
Adaptive Behavior Scale (r=0.479, p<0.05). No other associations were detected.
CONCLUS IONS: Our results indicate that elevated urine levels of the endogeneous
psychotomimetic molecule bufotenine may play a role in ASD and schizophrenia,
and can be correlated with hyperactivity scores in autism.
102
Copyright © 2010 Neuroendocrinology Letters ISSN 0172–780X • www.nel.edu
Enzo Emanuele, Roberto Colombo, Valentina Martinelli, Natascia Brondino, Mara Marini, Marianna Boso, Francesco Barale, Pierluigi Politi
INTRODUCTION
Previous studies have suggested that endogenous
psychotomimetic molecules may be involved in the
pathogenesis of major psychiatric disorders (Osmond
& Smythies 1952; Smythies 1983; Ciprian-Ollivier et al.
1988). Bufotenine (5-hydroxy-N,N-dimetyltryptamine)
is a tryptamine alkaloid derived from the double meth-
ylation of serotonin commonly found in a number of
mammals and in several amphibian groups (Takeda
1994). There is evidence to suggest that bufotenine has
potent psychotropic actions in humans, probably due to
its similar physiological and structural features to LSD
on the 5HT2 receptor (McBride 2000). Interestingly, an
endogenous production of bufotenine has been previ-
ously reported in patients with several psychiatric dis-
ease, and urine levels of this molecule have been found
to be elevated in subjects with schizophrenia in numer-
ous (Fischer & Spatz 1968; Räisänen et al. 1984; Fischer
et al. 1971; Narasimhachari & Himwich 1972; Cottrell et
al. 1977) but not all (Perry et al. 1966; Siegel 1956; Rod-
night 1956; Wyatt et al. 1973) studies. Of interest is also
the observation that bufotenine has been reported to be
higher in the urine of patients with autistic spectrum
disorder (ASD) (Piggott 1979; Himwich et al. 1972)
as well as their parents (Narasimhachari & Himwich
1975). Notably, a previous study has detected bufotenine
at significant amounts in 32/47 autistic patients and in
18/18 patients with mental retardation, whereas it was
found in only 2 out of 200 controls (Takeda et al. 1995).
Urinary excretion of bufotenine has been also found to
be higher in drug-free violent offenders, and levels of
this molecule have been found to positively associated
with suspiciousness and aggression and negatively with
socialization scores (Kärkkäinen et al. 1995; Räisänen
et al. 1984). In this study, we assessed urine levels of
bufotenine in subjects with autistic spectrum disorder
(ASD), schizophrenia and healthy comparison subjects
free of psychiatric symptoms. We also sought to investi-
gate whether urine bufotenine levels may be associated
with the clinical characteristics of psychiatric patients.
MATERIALS AND METHODS
Subjects
We measured urine bufotenine levels in the following
three groups of subjects: adults with severe ASD (n =15;
13 males and 2 females, mean age: 31.0 ± 7.5 years),
patients with schizophrenia (n=15, 10 males and 5
females, mean age: 32.7 ± 8.6 years), and normal con-
trol subjects (n=18; 15 males and 3 females, mean age
31.9 ± 8.0 years). Patients with ASD were recruited from
a single farm community center specifically designed
for individuals with autism (Cascina Rossago, San
Ponzo Semola, Pavia, Italy). The diagnosis of ASD was
confirmed in all participants jointly by two indepen-
dent psychiatrists specializing in autism who made the
diagnosis according to the guidelines of the Structured
Clinical Interview for Axis I DSM-IV Disorders, Clini-
cal Version (First et al. 1996). All patients in the present
study scored more than 30 on the Childhood Autism
Rating Scale (CARS) (Schopler et al. 1988), the standard
threshold used to distinguish autism. The Vineland
Adaptive Behavior Scale (Sparrow et al. 1984) was used
to measure adaptive behaviors in ASD individuals.
Patients with schizophrenia were recruited from
the Department of Psychiatry at the Pavia University
School of Medicine and from Outpatients Departments
from which we receive referrals. Each patient was given
a diagnostic assessment by an experienced psychiatrist
based on the Structured Clinical Interview for DSM-IV
(American Psychiatric Association 2005). The psycho-
pathological status of patients with schizophrenia was
assessed using the Brief Psychiatric Rating Scale (BPRS)
(Ventura et al. 2000). All psychiatric patients were either
medication-naive (first-onset) or medication-free for at
least four months. Control subjects were recruited from
healthy blood donors or volunteers who had helped in
other studies within our institutions. Comparison sub-
jects were drawn from the same geographical area as
our patient group, aiming to recover the basic demo-
graphics of the regions from which the patients were
recruited. All controls had no past or present history
of any psychiatric disease and none of them had ever
taken medications for psychiatric conditions. Addition-
ally, subjects with axis-I diagnosis of first degree rela-
tives were not included in this group.
The study was approved by the local ethics com-
mittee in accordance to the Helsinki Declaration and
written informed consent was obtained from all partici-
pants or legal guardians.
Laboratory methods
Specimens of the middle urinary flow were collected
using a standard sterile urine container, transported
in the dark in a refrigerated bag and frozen at –40 °C
within 3 hours of collection. Bufotenine concentrations
in urine were determined using a high-performance
liquid chromatography-mass spectrometry (HPLC-
MS) method as previously described (Kärkkäinen et al.
2005), with slight modifications. Urine bufotenine was
determined in a quality control sample with within-
series and between-series coefficients of variation of
3.8 and 4.6%, respectively. Since laboratory personnel
were blinded to the clinical status of the study partici-
pants, any possible measurement error was likely to be
non-differential.
Data analysis
Continuous variables were tested for normal distribu-
tion with the Kolmogorov–Smirnov statistics. Since all
variables were normally distributed only parametric
statistics were used. Continuous data are expressed as
means ± standard deviations. Comparison of categori-
cal variables was generated by the χ2 test. For three
group comparisons of quantitative variables, a one-way
103
Neuroendocrinology Letters Vol. 31 No. 1 2010 • Article available online: http://node.nel.edu
Elevated urine levels of bufotenine in patients with autistic spectrum disorders and schizophrenia
analysis of variance (ANOVA) with post-hoc Dunn’s
testing was performed. Correlations among variables
were computed with the use of Pearson’s correlation
coefficients. All statistical analyses were carried out
using SPSS version 16.0 (SPSS Inc., Chicago, IL, USA)
and GraphPad Prism version 4.0 (GraphPad Software
Inc., San Diego, CA, USA). A two-tailed p-value <0.05
was considered statistically significant.
RESULTS
The general characteristics of the study partici-
pants are depicted in Table 1. No significant differ-
ences in demographic and clinical parameters were
detected among different groups. Urine bufotenine
levels were significantly higher in the ASD group
(3.30 ± 0.49 g/L, p<0.05) and in patients with schizo-
phrenia (4.39 ± 0.43 g/L, p<0.001) compared with
controls (1.53 ± 0.30 g/L, Figure 1). Although there
was a trend towards higher levels of urine bufotenine
in patients with schizophrenia than in ASD, this differ-
ence failed to reach the statistical significance thresh-
old. Among patients with ASD, there was a significant
positive correlation between urine bufotenine and
hyperactivity scores on the Vineland Adaptive Behav-
ior Scales (r=0.479, p<0.05). No significant correlations
were found between BPRS scores and urine bufotenine
levels in patients with schizophrenia (data not shown).
DISCUSSION
The present study provides evidence of a significant ele-
vation of urine bufotenine levels in patients with ASD
and schizophrenia compared with a matched control
population. Additionally, concentrations of bufotenine
in urine were found to be positively associated with
hyperactivity scores on the Vineland Adaptive Behav-
ior Scales in ASD subjects. Although no correlation was
evident between urinary levels of this molecule and the
severity of schizophrenia symptoms, our data point to a
subtle but definite role of this endogenous pyschotomi-
metic molecule in ASD and schizophrenia.
Subtle alterations in serotonin metabolism have
been suggested to occur and play a role in the patho-
genesis of schizophrenia and autism. Bufotenine has
been found to act as a potent endogenous hallucino-
genic factor, with an activity similar to LSD at the pur-
ported hallucinogenic serotonin receptors, 5HT2A and
Tab. 1. General characteristics of the study participants.
ASD patients
(n=15)
Schizophrenia
(n=15)
Controls
(n=18) p-value
Age, years 31.0 ± 7.5 32.7 ± 8.6 31.9 ± 8.0 0.59
Males/females 13/2 10/5 15/3 0.35
Body mass index, kg/m2 23.9 ± 3.3 24.7 ± 3.6 24.6 ± 3.4 0.47
Glucose, mmol/L 4.6 ± 0.7 4.5 ± 0.6 4.7 ± 0.7 0.77
Creatinine, mg/dL 0.82 ± 0.14 0.84 ± 0.18 0.83 ± 0.19 0.61
Total cholesterol, mmol/L 4.6 ± 0.7 4.8 ± 0.8 4.7 ± 0.7 0.69
LDL cholesterol, mmol/L 2.8 ± 0.8 2.9 ± 1.0 3.1 ± 1.0 0.30
HDL cholesterol, mmol/L 1.4 ± 0.4 1.5 ± 0.4 1.4 ± 0.5 0.89
Triglycerides, mmol/L 1.6 ± 0.9 1.6 ± 1.0 1.5 ± 1.1 0.67
Serum sodium, mmol/L 141.9 ± 3.2 142.0 ± 3.5 140.8 ± 3.0 0.91
Serum potassium, mmol/L 4.1 ± 0.5 4.0 ± 0.5 4.0 ± 0.6 0.84
Systolic blood pressure, mmHg 128 ± 9 130 ± 10 126 ± 7 0.21
Diastolic blood pressure, mmHg 82 ± 7 84 ± 11 81 ± 6 0.18
Urine bufotenine, μg/L 3.30 ± 0.49 4.39 ± 0.43 1.53 ± 0.30 < 0.001
p-values are calculated by means of ANOVA or χ2 test, as appropriate.
Bufotenin [μg/L]
autistics controls schizophrenics
0
2
4
6
8
Fig. 1. Scatter diagram for urine bufotenine in the three study
groups (ASD, controls, and schizophrenia). Horizontal lines
across the scatter diagram represent mean values.
104
Copyright © 2010 Neuroendocrinology Letters ISSN 0172–780X • www.nel.edu
Enzo Emanuele, Roberto Colombo, Valentina Martinelli, Natascia Brondino, Mara Marini, Marianna Boso, Francesco Barale, Pierluigi Politi
5HT2C (McBride 2000). Although some reports have
reported detectable levels of urinary bufotenine in sev-
eral groups of patients with major psychiatric illnesses
(Fischer & Spatz 1968; Räisänen et al. 1984; Fischer et
al. 1971; Narasimhachari & Himwich 1972; Cottrell
et al. 1977), other authors were unable to confirm the
presence of this molecule (Perry et al. 1966; Siegel 1956;
Rodnight 1956; Wyatt et al. 1973). One of the reasons
for such discrepancy may be the lack of standardization
of methods for the determination of urinary bufoten-
ine levels and different ways in which urine samples are
handled. In this study, we used an HPLC-MS method to
detect urine bufotenine as previously described (Kärk-
käinen et al. 2005). Of note, we were able to confirm the
presence of a detectable amounts of bufotenine not only
in patients with psychiatric disorders, but also in con-
trols. This result is in keeping with those obtained by
Kärkkäinen and coworkers (Kärkkäinen et al. 2005), but
conflict with those obtained by earlier studies (Perry et
al. 1966; Siegel 1956; Rodnight 1956; Wyatt et al. 1973).
An improved sensitivity of the laboratory techniques
used to assess urine bufotenine may account for such
findings.
In our study, we found that urine bufotenine was sig-
nificantly higher in patients with ASD and schizophre-
nia compared with controls. These findings were in line
with those obtained by previous studies in schizophre-
nia and autism (Fischer & Spatz 1968; Räisänen 1984;
Fischer et al. 1971; Narasimhachari & Himwich 1972;
Cottrell et al. 1977, Himwich et al. 1972; Piggott 1979).
However, previous studies did not clarify whether urine
bufotenine might be associated with the clinical features
of major psychiatric illnesses. In our report, a signifi-
cant positive correlation between hyperactivity scores
and urine bufotenine levels was evident in patients with
ASD. It is feasible to hypothesize that increased levels of
bufotenin in autistic subjects with hyperactivity can be
a biological correlate of their behavior or – more specu-
latively – be a causal factor. Intriguingly, our data paral-
lel those of Kärkkäinen et al. (1995) who showed that
violent offenders with paranoid personality traits have
higher urinary levels of bufotenine than other violent
offenders. Additionally, Räisänen et al. have suggested
that urinary excretion of bufotenine is increased in
violent offenders with paranoid symptoms and family
violence (Räisänen et al. 1984). Although these findings
may prompt intriguing hypotheses on the possible asso-
ciation of bufotenine levels in urine and aberrant behav-
iors, further investigations are needed in larger sample
size to draw a more definite conclusion on this issue.
Caveats of this study merit consideration. The chief
limitations of our report are the small sample size and
its cross-sectional design. We thus observed associa-
tions, not prediction or causation. Therefore, it remains
to be established whether increased bufotenine level
observed in autism is a cause or a consequence of dis-
ease. In addition, we limited our analysis to bufotenine
levels as measured in the urine. In light of this limita-
tion, it remains to be established whether the increase
of peripheral bufotenine observed in autism may be
paralleled by similar changes in the central nervous
system. In this regard, it has been previously suggested
that the CNS is a favored site for the accumulation of
bufotenine, probably because of the lipophilic propteri-
ties of this substance and the slow catabolism of methyl-
ated indolamines in the brain (Kärkkäinen et al. 2005).
The presence of a significantly higher urinary bufoten-
ine concentration is another evidence of the complex
serotoninergic alterations occurring in autism and
schizophrenia, which probably includes an abnormal
methylation of this neurotransmitter.
In summary, our results indicate that elevated urine
bufotenine levels may play a role in schizophrenia and
ASD and may be correlated with hyperactivity scores
in autism.
ACKNOWLEDGEMENTS
The excellent nursing assistance of Roberto Albasi is
gratefully acknowledged.
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