Combination of experimental and in silico methods for the assessment of the phototransformation products of the antipsychotic drug/metabolite Mesoridazine

DOI: 10.1016/j.scitotenv.2017.08.040

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Science of the Total Environment p. 697 - 711 (2018)

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Wilde, Marcelo L.

Menz, Jakob

Leder, Christoph

Kümmerer, Klaus

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Article abstract of DOI:10.1016/j.scitotenv.2017.08.040

The lack of studies on the fate and effects of drug metabolites in the environment is of concern. As their parent compounds, metabolites enter the aquatic environment and are subject to biotic and abiotic process. In this regard, photolysis plays an important role. This study combined experimental and in silico quantitative structure-activity relationship (QSAR) methods to assess the fate and effects of Mesoridazine (MESO), a pharmacologically active human drug and metabolite of the antipsychotic agent Thioridazine, and its transformation products (TPs) formed through a Xenon lamp irradiation. After 256 min, the photodegradation of MESO ? besylate (50 mg L^{? 1}) achieved 90.4% and 6.9% of primary elimination and mineralization, respectively. The photon flux emitted by the lamp (200–600 nm) was 169.55 J cm^{? 2}. Sixteen TPs were detected by means of liquid chromatography-high resolution mass spectrometry (LC-HRMS), and the structures were proposed based on MSⁿ fragmentation patterns. The main transformation reactions were sulfoxidation, hydroxylation, dehydrogenation, and sulfoxide elimination. A back-transformation of MESO to Thioridazine was evidenced. Aerobic biodegradation tests (OECD 301 D and 301F) were applied to MESO and the mixture of TPs present after 256 min of photolysis. Most of TPs were not biodegraded, demonstrating their tendency to persist in aquatic environments. The ecotoxicity towards Vibrio fischeri showed a decrease in toxicity during the photolysis process. The in silico QSAR tools QSARINS and US-EPA PBT profiler were applied for the screening of TPs with character of persistence, bioaccumulation, and toxicity (PBT). They have revealed the carbazole derivatives TP 355 and TP 337 as PBT/vPvB (very persistent and very bioaccumulative) compounds. In silico QSAR predictions for mutagenicity and genotoxicity provided by CASE Ultra and Leadscope indicated positive alerts for mutagenicity on TP 355 and TP 337. Further studies regarding the carbazole derivative TPs should be considered to confirm their hazardous character.

Full text of DOI:10.1016/j.scitotenv.2017.08.040

STOTEN-23604; No of Pages 15

Science of the Total Environment xxx (2017) xxx –xxx

Contents lists available at ScienceDirect

Science of the Total Environment

journal homepage: w ww.elsevier.com/locate/scitotenv

Combination of experimental and in silico methods for the assessment of

the phototransformation products of the antipsychotic drug/

metabolite Mesoridazine

Marcelo L. Wilde ^a, Jakob Menz ^b, Christoph Leder ^b, Klaus Kümmerer ^b,

⁎

Formerly: Sustainable Chemistry and Material Resources, Institute of Sustainable Environmental Chemistry, Leuphana University Lüneburg, C13, DE-21335 Lüneburg, Germany

Sustainable Chemistry and Material Resources, Institute of Sustainable Environmental Chemistry, Leuphana University Lüneburg, C13, DE-21335 Lüneburg, Germany

H I G H L I G H T S

G R A P H I C A L A B S T R A C T

• Photolysis of the antipsychotic drug/me-

tabolite MESO was simulated through

Xe lamp.

• Sixteen TPs were detected and elucidat-

ed by means of an UHPLC-HRMSⁿmeth-

od.

• MESO and TPs were not readily biode-

gradable according to OECD 301D and

301F tests.

• Toxicity in bacteria decreased during

photolysis proportionally to the concen-

tration of MESO.

• In silico QSAR predicted the carbazole

derivative TPs as PBT/vPvB and indicat-

ed positive alerts for mutagenicity.

a r t i c l e i n f o

a b s t r a c t

Article history:

The lack of studies on the fate and effects of drug metabolites in the environment is of concern. As their parent

compounds, metabolites enter the aquatic environment and are subject to biotic and abiotic process. In this re-

gard, photolysis plays an important role. This study combined experimental and in silico quantitative struc-

ture-activity relationship (QSAR) methods to assess the fate and effects of Mesoridazine (MESO), a

pharmacologically active human drug and metabolite of the antipsychotic agent Thioridazine, and its transforma-

tion products (TPs) formed through a Xenon lamp irradiation. After 256 min, the photodegradation of MESO⋅

besylate (50 mg L⁻¹) achieved 90.4% and 6.9% of primary elimination and mineralization, respectively. The pho-

ton ﬂux emitted by the lamp (200–600 nm) was 169.55 J cm⁻². Sixteen TPs were detected by means of liquid

chromatography-high resolution mass spectrometry (LC-HRMS), and the structures were proposed based on

MSⁿfragmentation patterns. The main transformation reactions were sulfoxidation, hydroxylation, dehydroge-

nation, and sulfoxide elimination. A back-transformation of MESO to Thioridazine was evidenced. Aerobic bio-

degradation tests (OECD 301 D and 301F) were applied to MESO and the mixture of TPs present after 256 min

of photolysis. Most of TPs were not biodegraded, demonstrating their tendency to persist in aquatic environ-

ments. The ecotoxicity towards Vibrio ﬁscheri showed a decrease in toxicity during the photolysis process. The

in silico QSAR tools QSARINS and US-EPA PBT proﬁler were applied for the screening of TPs with character of per-

sistence, bioaccumulation, and toxicity (PBT). They have revealed the carbazole derivatives TP 355 and TP 337 as

PBT/vPvB (very persistent and very bioaccumulative) compounds. In silico QSAR predictions for mutagenicity and

Received 13 June 2017

Received in revised form 3 August 2017

Accepted 4 August 2017

Available online xxxx

Editor: D. Barcelo

Keywords:

Metabolite

Transformation products

Ready biodegradability

Ecotoxicity

QSAR

PBT/vPvB

⁎

Corresponding author at: Nachhaltige Chemie und Stofﬂiche Ressourcen, Institut für Nachhaltige Chemie und Umweltchemie, Fakultät für Nachhaltigkeit, Leuphana Universität

Lüneburg, Scharnhorststraße 1/C13, D-21335 Lüneburg, Germany.

E-mail addresses: marcelolw@gmail.com (M.L. Wilde), jakob.menz@uni.leuphana.de (J. Menz), cleder@leuphana.de (C. Leder), klaus.kuemmerer@uni.leuphana.de (K. Kümmerer).

https://doi.org/10.1016/j.scitotenv.2017.08.040

Please cite this article as: Wilde, M.L., et al., Combination of experimental and in silico methods for the assessment of the phototransformation

products of the antipsychotic drug/metabolite Mesoridazine, Sci Total Environ (2017), https://doi.org/10.1016/j.scitotenv.2017.08.040

M.L. Wilde et al. / Science of the Total Environment xxx (2017) xxx–xxx

genotoxicity provided by CASE Ultra and Leadscope® indicated positive alerts for mutagenicity on TP 355 and TP

337. Further studies regarding the carbazole derivative TPs should be considered to conﬁrm their hazardous

character.

1. Introduction

effects have been found such as cardiac toxicity and QT-prolongation,

which has led to a withdrawal in some countries (Jin et al., 2014). On

The lack of studies on the fate and effects of drug metabolites in the

environment is of great concern. The metabolism of drugs in humans in-

volves typically the transformation of the parent drug into more polar

and consequently more soluble compounds, which facilitate the drug

elimination (Celiz et al., 2009; Santos et al., 2010). The presence of

human metabolites has been demonstrated in environmental compart-

ments and some of them in same concentration levels as the parent

compounds (Langford and Thomas, 2011; López-Serna et al., 2013;

Osorio et al., 2014).

Furthermore, once pharmaceuticals and their metabolites achieve

wastewater treatment plants and surface waters, they are subject to

natural attenuation processes, i.e. biotic and abiotic transformations

(Zhu et al., 2015; Zonja et al., 2016). Biotic transformations are normally

carried out by organisms such as bacteria and fungi leading to its com-

plete mineralization or the formation of unknown transformation prod-

ucts (TPs) (Fatta-Kassinos et al., 2011; Kümmerer, 2010, 2009).

Photolysis is an important abiotic process and plays a major role in the

environmental fate of pharmaceuticals and their metabolites. In general,

pharmaceuticals and metabolites possess structural moieties, which are

able to absorb sunlight radiation leading to direct and indirect (sensi-

tized) photolysis (Du et al., 2014; Fatta-Kassinos et al., 2011; Lin et al.,

2013).

An increase in toxicity due to the formation of a more toxic cocktail

of TPs by photolysis has been reported in several studies (Wang and Lin,

2014; Mahmoud et al., 2014; Herrmann et al., 2015). This chronic expo-

sure to residues of pharmaceutical, metabolites, and TPs can generate

subtle effects on aquatic organisms and in a risk to human health by

consumption of contaminated waters and even through contaminated

food (Paltiel et al., 2016; Vandermeersch et al., 2015; Wu et al., 2014).

Many human metabolites and TPs still contain structural similarities

to the parent compounds and present pharmacophore groups attaining

similar activity (Cwiertny et al., 2014; Fura, 2006). Besides,

photodegradation might not entirely change the pharmacophore struc-

ture of the parent compound. Zhu et al. (2015) pointed out that the

photodegradation of amlodipine (AML), diltiazem (DIL), and verapamil

(VER) under UV (254 nm) irradiation led to TPs still containing intact

pharmacophore structures. The TPs of VER did not present ecological

risks. Conversely, the photodegradation of AML and DIL under typical

of disinfection UV ﬂuence led to TPs containing pharmacophores,

which could present potential risks. Therefore, photodegradation of

pharmaceuticals might not entirely eliminate the risk of pharmaceuti-

cals in the environment.

the other hand, Thioridazine is still used in antipsychotic therapy and

has been found as a potent antimicrobial agent (Amaral, 2012;

Thanacoody, 2007, 2011). This could lead to an elevated environmental

occurrence of MESO and possible resulting TPs. The photostability of

MESO under UV light of 254 nm and 366 nm was tested in methanolic

solutions and epimerization of was observed (De Gaitani et al., 2004),

but no extensive study on the environmental fate and effects of the pho-

tolysis of MESO has been reported yet in the literature. Besides, MESO

was also identiﬁed as a TP of the parent compound Thioridazine in our

previous studies of photolysis and Fenton process (Wilde et al., 2017,

2016).

The assessment of the photolytic fate of human metabolites and

their TPs is even more scarce in the literature (Bonvin et al., 2013).

Therefore, the aim of the present study was to assess the fate and effects

of the TPs formed from the antipsychotic and human metabolite MESO

through a Xenon lamp irradiation. The elucidation and proposal of the

chemical structure of TPs were carried out by means of ultra-high per-

formance liquid chromatography tandem high-resolution Orbitrap

mass spectrometry (UHPLC-HRMS). The fate and effects of MESO and ir-

radiated solutions were tested experimentally by means of ready biode-

gradability according to OECD 301D and 301F tests, and the toxicity was

tested towards luminescent bacteria (Vibrio ﬁscheri). In silico models

based on quantitative structure-activity relationships (QSAR) were ap-

plied to screen for TPs with the character of persistence, bioaccumula-

tion, and toxicity (PBT). Furthermore, QSAR models were also

employed for the prediction of genotoxic and mutagenic activities of

identiﬁed TPs.

2. Materials and methods

2.1. Chemicals

Mesoridazine besylate (MESO⋅besylate, CAS Nr. 32672-69-8) was

acquired from Santa Cruz Biotechnology (Dallas, Texas, USA). 3,5-

Dichlorophenol (97%, CAS Nr. 591-35-5), Chloramphenicol (98%, CAS

Nr. 56-75-7) were purchased from Sigma-Aldrich (Deisenhofen,

Germany). Benzenesulfonic acid sodium salt (i.e. besylate) (98%, CAS

Nr. 515-42-4) was acquired from Acros Organics (New Jersey, USA). Or-

ganic solvents were of LC-MS grade and provided by VWR (Darmstadt,

Germany). Aqueous solutions were prepared in ultrapure water

(Q1:16.6 MΩ⋅cm and Q2:18.2 MΩ⋅cm, Ultra Clear UV TM, Barsbüttel,

Germany). All other chemicals used were of recognized analytical grade.

Many drug metabolites have been studied and identiﬁed within

pharmaceutical drug development and the ones that retained a phar-

macological activity have often attained importance as pharmaceuticals.

Mesoridazine (MESO, or Thioridazine-2-sulfoxide) is such an example.

It is a phenothiazine-derived antipsychotic agent and a major metabo-

lite of Thioridazine in humans and animals. MESO is formed through a

sulfoxidation on the R2-substituent (SCH₃) at the C2-position of the

phenothiazine tricyclic ring. In the human body, approximately 30% of

orally administered THI is excreted in the urine and 50% of the original

dose is excreted in the feces (Eiduson and Geller, 1963). The mean

total excretion of MESO in the urine of humans is 6.3% (Hawesx, 1993).

Owing to a higher antipsychotic activity than Thioridazine (Obach,

2013; Ravyn et al., 2013), MESO was proposed for treatment of psychot-

ic disease such as schizophrenia. Nevertheless, some unwanted side

2.2. Photolysis setup

The photolysis experiments were carried out in a 1000 mL cylindri-

cal immersion-type batch reactor. The irradiation was simulated by

means of a UV/VIS 150 W Xenon lamp (TXE 150 W, UV Consulting

Peschl, Mainz, Germany) surrounded by an ilmasil quartz glass used

as a cooling jacket separating the lamp from the solution immersed

into a 800 mL of synthetic solution of MESO diluted in ultrapure water

(Text S1, Supplementary material). The irradiation of the lamp, mea-

sured with a Black Comet UV-VIS spectroradiometer model C

(StellarNet Inc., Florida, USA), was: 200–280 nm: 1.01 W m⁻²

;

280–315 nm: 3.29 W m⁻²; 315–380 nm: 12.91 W m⁻², 380–850 nm:

243.16 W m⁻². The spectrum of the lamp and the molar extinction

Please cite this article as: Wilde, M.L., et al., Combination of experimental and in silico methods for the assessment of the phototransformation

products of the antipsychotic drug/metabolite Mesoridazine, Sci Total Environ (2017), https://doi.org/10.1016/j.scitotenv.2017.08.040

M.L. Wilde et al. / Science of the Total Environment xxx (2017) xxx–xxx

kinetic model was proposed to ﬁt the exponential decay of MESO ac-

cording to Eq. (1):

_obst

C ¼ C₀∙e^−k

ð1Þ

where C is the concentration of MESO during the photolysis, C₀is the

initial concentration of MESO, k_obsis the observed kinetic constant, t is

irradiation time. The half-life (t ) was calculated according to Eq. (2):

ꢀ

0:693

t₁

ð2Þ

k_obs

The kinetics of the experimental data were ﬁtted with the software

SigmaPlot 12 (Systat Software, USA) by means of nonlinear model ﬁt re-

gressions. The statistical analysis of the ﬁtting was performed by means

of ANOVA.

The quantum yield for MESO was calculated using the modiﬁed

equation for polychromatic irradiation sources (Eq. (3)) (Zepp, 1978;

Zepp and Cline, 1977).

NPOC (MESO Bes (50 mg L ))

NPOC (MESO Bes (70.46 mg L ))

25 50 75 100 125 150 175 200 225 250 275

Time (min)

k_obs

ꢁ ꢂ

Φ_MESO

ð3Þ

200

2:303 ꢀ ∑ I_0λϵ_λꢀ l ꢀ

600

where ∅_MESOis the apparent quantum yield (mol Einstein⁻¹) of MESO,

MESO Besylate

Xe Lamp (TXE 150 W)

k_obsis the kinetic constant of primary elimination (s⁻¹). The term

λ_n

∑

I_0λϵ_λis the sum of the spectral superposition of the irradiation

λ_i

emitted by the lamp on the target compound at a determined wave-

length range. Thus, I_0λis the spectral photon ﬂux rate at each wave-

length from 200 to 850 nm (Einstein m⁻²s⁻¹), ϵ_λis the molar

extinction coefﬁcient of MESO (m⁻²mol⁻¹), l is the radiation pathway

from the lamp to solution bulk (0.015 m), A is area of reactor (0.062 m²),

and V is the volume of reactor (0.0011 m³).

The spectral photon ﬂux rate of the lamp was calculated according to

Eq. (4).

E∙λ∙5:03∙10¹⁵

I_λ

ð4Þ

N_A

where E is the irradiance of the lamp (W m⁻²), λ is the wavelength (m),

N_Ais Avogadro's number (6.022

10²³mol⁻¹), and 5.03

× 10¹⁵m⁻²s⁻¹is a constant.

Wavelenght (nm)

2.3. Instrumental analysis

Fig. 1. (A) Non-purgeable organic carbon (NPOC) removal and primary elimination of

Mesoridazine (MESO) through photolysis by means of Xe lamp (TXE 150 W) irradiation.

Initial conditions: [MESO⋅besylate] 50 mg L⁻¹(35.48 mg L⁻¹of MESO only), pH 7.0,

temp.: 20 2 °C (n = 2); [MESO·besylate] 70.46 mg L⁻¹(50.00 mg L⁻¹of MESO only),

pH 7.0, temp.: 20

150 W) and molar absorption coefﬁcient of MESO⋅besylate (ε_PPL).

The primary elimination was measured by HPLC-DAD-FLD Promi-

nence HPLC (Shimadzu, Duisburg, Germany) according to the modiﬁed

method of Trautwein and Kümmerer (2012a). A non-target approach

was performed for the detection and elucidation of TPs using an Agilent

Technologies HPLC 1100 series (Agilent Technologies, Böblingen,

Germany) tandem Mass Spectrometer Esquire 6000^plusIon Trap with

atmospheric pressure electrospray ionization (ESI) interface (Bruker

Daltonics GmbH, Bremen, Germany) in positive ion mode (LC-ESI-IT-

MSⁿ). A more accurate elucidation of the TPs was proposed by using a

Dionex Ultimate 3000 UHPLC system (Dionex, Idstein, Germany) tan-

dem to an LTQ Orbitrap-XL high-resolution mass spectrometer

(HRMS) with H-ESI ion source (Thermo Scientiﬁc, Bremen, Germany).

The chromatographic separation in both LC-MS/MS instruments was

carried out on a reverse phase column C18 ec RP18 CC 125-2 mm

Nucleodur 100-3 and guard column RP18 CC 8–2 mm Nucleodur 100-

3 (Macherey-Nagel, Düren, Germany). Detailed information the chro-

matographic methods can be found in Text S2 (Supplementary

material).

2 °C (n = 1). (B) Irradiance (W m⁻²) of the Xenon lamp (TXE

coefﬁcient of MESO are depicted in Fig. 1 (B). The initial concentration of

MESO⋅besylate used in the experiments were 50 mg L⁻¹(correspond-

ing to 35.48 mg L⁻¹of MESO) and 70.46 mg L⁻¹(50.00 mg L⁻¹of

MESO). They were chosen in order to allow the detection of as many

TPs as possible and further experimental evaluation of aerobic biodegra-

dation and toxicity tests, respectively. The experiments were carried out

at pH 7 and no adjustments in pH of the solution were carried out dur-

ing and after the experiments. The temperature was held at 20 2 °C

with a circulating cooler (WKL230, LAUDA, Berlin, Germany).

2.2.1. Kinetics of degradation and quantum yield calculations

The mineralization was determined by monitoring the non-

purgeable organic carbon (NPOC) through a total organic carbon ana-

lyzer (TOC-Vcpn, Shimadzu GmbH, Duisburg, Germany) with ASI-V

auto-sampler.

In general, a photochemical reaction does not follow a speciﬁc reac-

tion order, but the kinetic behavior dependent on the absorption condi-

tions of the target compound (Oppenländer, 2003). Thus, a ﬁrst-order

Please cite this article as: Wilde, M.L., et al., Combination of experimental and in silico methods for the assessment of the phototransformation

products of the antipsychotic drug/metabolite Mesoridazine, Sci Total Environ (2017), https://doi.org/10.1016/j.scitotenv.2017.08.040

M.L. Wilde et al. / Science of the Total Environment xxx (2017) xxx–xxx

2.4. Aerobic biodegradation tests

3. Results and discussion

MESO⋅besylate and the mixture resulting after 256 min of

photodegradation were submitted to Closed Bottle Test (CBT) (OECD

301 D) and Manometric Respirometry Test (MRT) (OECD 301F) accord-

ing to the OECD guidelines (OECD, 1992). The CBT was carried out by

3.1. Photolysis

The photon ﬂux emitted by the lamp (wavelength range 200–

600 nm) during 256 min of irradiation was 169.55 J cm⁻². Fig. 1

(A) shows that the photolysis achieved 90.4% ( 7.9%) of primary elim-

ination and 6.9% ( 4.2%) of mineralization after 256 min for the initial

concentration of 50 mg L⁻¹MESO⋅besylate (35.48 mg L⁻¹of MESO).

For the initial concentration of 70.46 mg L⁻¹MESO⋅besylate

(50.00 mg L⁻¹of MESO), the photolysis reached after 256 min of pho-

tolysis was 70.8% of primary elimination and only 2.75% of mineraliza-

using low nutrient load and low bacteria density (10²–10⁵CFU mL⁻¹

)

(OECD, 1992), in the dark at 20 ( 1) °C for a period of 28 days. The ox-

ygen demand (OD) was measured by a sensor Fibox 3 (Presens, Regens-

burg, Germany) (Friedrich et al., 2013). The MRT was performed by

using an OxiTop® OC-110 system (WTW, Weilheim, Germany) under

constant stirring in the dark during 28 days at 20 ( 1) °C and using

the same mineral medium as for CBT (OECD, 1992). The inoculum

used was a sample of efﬂuent from the municipal STP Lüneburg

(Abwasser, Grün & Lüneburger Service GmbH (AGL), Lüneburg,

Germany), which serves a regional population equivalent of 144,000 in-

habitants. Further information can be found in Text S3 (Supplementary

material). The peak area analysis of MESO and TPs was carried out at day

0 and after 28 days by LC-ESI-IT-MSⁿ(see Section 2.3).

tion. These results indicated that the photolysis was

a mere

transformation process, reinforcing the need to measure mineralization

in photolysis experiments and to include results from the assessment of

fate and effects of TPs in the environment.

As depicted in Fig. 1 (A), the NPOC removal was ﬁtted with the zero-

order kinetic model with k_DOC3.43 × 10⁻⁶mol L⁻¹s⁻¹(95% conﬁ-

obs

dence interval (CI): [2.29 × 10⁻⁶− 4.57 × 10⁻⁶]) and r²0.784 (p b

0.05) for the initial concentration of 50.00 mg L⁻¹MESO⋅besylate

(29.76 mg L⁻¹of organic C). The zero-order kinetic constant for the ini-

tial concentration of 70.46 mg L⁻¹MESO⋅besylate (41.94 mg L⁻¹of or-

2.5. Assessment of the toxicity by modiﬁed luminescent bacteria test

The toxicity on bacteria was assessed in a modiﬁed luminescent bac-

teria test (LBT) with Vibrio ﬁscheri NRRL-B-11177 (Hach-Lange GmbH,

Düsseldorf, Germany) according to (Menz et al., 2013). This bioassay

was specially designed for the combined determination of short-term

(30 min) and long-term (24 h) inhibition of bacterial bioluminescence.

In addition, the inhibition of bacterial growth was evaluated at the tran-

sition to the stationary phase after 14 h incubation. Detailed information

about the experimental procedure of the LBT can be found in Text S4

(Supplementary material). Inhibitory effects were determined in rela-

tion to a nonexposed control. A minimum threshold of 20% inhibition

was applied for the identiﬁcation of signiﬁcant effects. The ﬁtting of

concentration-response curves was done with the software package

SigmaPlot 11 (Systat Software) using a four-parameter Hill model (Eq.

(5)).

ganic C) was k_DOC1.47 × 10⁻⁶mol L⁻¹s⁻¹(95% CI: [−3.76 × 10⁻⁷

−

3.31 × 10⁻⁶]) an^od^bsr²0.339 (p = 0.1018). The primary elimination of

MESO was ﬁtted by a ﬁrst-order exponential decay for both concentra-

tion studied as showed in Fig. 1 (A). The computed kinetic constant for

the photolysis of MESO (35.48 mg L⁻¹) was k_obs1.47 × 10⁻⁴s⁻¹(95%

CI: [1.35 × 10⁻⁴− 1.59 × 10⁻⁴]) with r²0.993 (p b 0.05), with a half-life

time (t ) of 4861.2 s. Considering MESO (50.00 mg L⁻¹) the k_obs8.39

× 10⁻⁵s⁻¹(95% CI: [7.95 × 10⁻⁵− 8.83 × 10⁻⁵]) with r²0.998 (p b

0.05) and the half-life time (t ) was 8179.2 s.

The apparent quantum yield of MESO (Φ_MESO) at pH 7 was estimated

to be 3.05 × 10⁻⁶

(

6.37 × 10⁻⁷) mol Einstein⁻¹(n = 2) using the ki-

netic slope method (Zepp, 1978; Zepp and Cline, 1977). At the pH 7, the

major species of MESO was calculated to be 93.95% of the tertiary amine

of the piperidine moiety protonated; whereas 6.05% are estimated to be

neutral (calculated using Marvin 6.0.3, 2013, (ChemAxon, http://www.

chemaxon.com). The monitoring of the pH during the photolysis

showed no perceptible changes in pH (see Text S6, Supplementary ma-

terial). The phototransformation of MESO occurs by either direct or in-

direct photolysis. In the direct photolysis, the absorbance of photons

plays an important role. The photolysis depends on both, the rate of

light absorption and the reaction quantum yield of the excited state.

As observed in Fig. 1 (B), the emission of the lamp overlaps the absor-

bance of MESO in the range of 280–400 nm.

ꢃ

ꢄ

−Hillslope

y ¼ min þ ð max− minÞ= 1 þ ðx=EC₅₀

ð5Þ

where min is the bottom of the curve, max is the top of the curve, Hillslope

is the slope of the curve at its midpoint and EC₅₀is the half-maximal effec-

tive concentration. The expected effect of MESO in the photolysis mixture

was predicted by inserting the measured concentration of MESO into the

plot equation of the ﬁtted concentration-response curve.

A hypothesized scheme concerning the possible pathways of MESO

through photolysis was proposed (Fig. 2). MESO can follow different

pathways under irradiation: (i) MESO in a competition with intersys-

tem crossing can undergo photoionization through direct photolysis

2.6. In silico quantitative structure-activity relationships (QSARs)

predictions

The neutral species (i.e. not considering protonated, deprotonated

and/or zwitterion microspecies) of the chemical structures investigated

were converted into the simpliﬁed molecular-input line-entry system

(SMILES) and subjected to in silico predictions. The QSARINS software

v.2.0 (Gramatica et al., 2014, 2013) based on the Insubria-PBT Index

(Papa and Gramatica, 2010) as implemented in the QSARINS-Chem

module of the software QSARINS and the US-EPA PBT Proﬁler v.2.000

(USEPA, 2012) were used in a consensus mode for the screening of

PBT compounds. Moreover, two other QSAR platforms were used for

the prediction of mutagenic and genotoxic activities: CASE Ultra (v.

1.6.0.3 MultiCASE Inc.) (Chakravarti et al., 2012; Saiakhov et al., 2013;

Saiakhov et al., 2014) and Leadscope® (v. 3.4.2-2) (Roberts et al.,

2000). The applied models included both statistical and rule-based sys-

tems as recommended by the ICH M7 guidelines (International

Conference on Harmonization (ICH), 2014) to predict mutagenicity

and were used for different endpoints as can be seen in Text S5 (Supple-

mentary material).

MESO^+•+ e⁻

h ν

iii

O₂

¹MESO*

h ν

MESO

viii

O₂

¹O₂

xii

−•

MESO^+•+ O₂

³MESO*

O₂

³O₂

O₂^−•+ 2H⁺

MESO^•+ H^•

Fig. 2. Hypothesized reaction scheme for the direct photolysis of Mesoridazine (MESO) in

aqueous solution in presence of natural dissolved oxygen. ROS: reactive oxygen species;

TPs: transformation products.

Please cite this article as: Wilde, M.L., et al., Combination of experimental and in silico methods for the assessment of the phototransformation

products of the antipsychotic drug/metabolite Mesoridazine, Sci Total Environ (2017), https://doi.org/10.1016/j.scitotenv.2017.08.040

M.L. Wilde et al. / Science of the Total Environment xxx (2017) xxx–xxx

forming MESO cation radical (MESO⁺•) and a hydrated electron

(Rodrigues et al., 2006), which in the presence of dissolved oxygen

can further form superoxide (O⁻₂•) (ix). However, direct photoioniza-

tion is unlikely to occur according to energetic considerations

(Fasnacht and Blough, 2002). Likewise, MESO is excited to a singlet

state (¹MESO*) (ii), where it can undergo intersystem crossing (IC) via

internal conversion to a triplet state (³MESO*) (iv) by a biphotonic or

monophotonic absorption process (Rodrigues et al., 2006), or, in the

presence of dissolved oxygen, it might form an MESO⁺• and O₂⁻• (viii).

The MESO⁺• can further form TPs (x) and reactive oxygen species

(ROS) (xi), or as proposed by Rodrigues et al. (2006) lead to the forma-

tion of MESO• and H• (vii). Alternatively, the singlet state ¹MESO* might

also undergo photoionization forming MESO⁺• and hydrated electron

(iii). Likewise, the triplet state ³MESO* can lead to the formation of

MESO• and H• through monophotonic homolytic cleavage (v),

MESO⁺• and O⁻₂• (vi), or even be quenched by triplet oxygen (xii)

returning to the ground state of MESO (Fasnacht and Blough, 2002).

The singlet oxygen in photolysis has been pointed out as a very im-

portant variable in the direct photolysis studies (Chen et al., 2008; Du

et al., 2014; Edhlund et al., 2006). Chen et al. (2008) have proposed

that oxygen singlet is formed through the quenching of triplet state of

MESO.

Sixteen TPs were found resulting from photolysis. Chromatographic

peaks with the same m/z [M + H]⁺and different retention times were

observed, indicating the formation of constitutional isomers. A possible

photolysis pathway based on these results is shown in Fig. 3. Nine peaks

with the m/z [M + H]⁺403 Da were found indicating sulfoxidation or

mono-hydroxylation. Based on the product ion of m/z 292.0461

(C₁₄H₁₄O₂NS₂) and 142.1224 (C₈H₁₆ON) indicating that the hydroxyl-

ation on TP 403 I might occur on the α-carbon of the R1-substituent

of MESO. TPs 403 II-V and VII have also presented the product ion of

m/z 142.1224 (C₈H₁₆ON) in the MS²and/or MS³spectra, but accordingly

to the fragmentation pattern the hydroxylation was proposed on the pi-

peridine moiety.

The exact position of the hydroxylation cannot be given based on the

MSⁿdata only. The fragmentation pattern of TPs 403 VI and VIII indicat-

ed that the hydroxylation happened at an aromatic ring of the tricyclic

phenothiazine ring. TP 403 IX was proposed to undergo a new

sulfoxidation on the R2-substituent forming Thioridazine-2-Sulfone

(Sulforidazine). The loss of the moiety S(O₂)CH₃evidenced by the prod-

uct ion of m/z 324.1656 (C₂₀H₂₄N₂S) supported this proposition.

Sulforidazine has been proposed as a metabolite of both Thioridazine

and Mesoridazine (Borges et al., 2007; De Gaitani et al., 2004; Eap

et al., 1996; Hawesx, 1993; Wójcikowski et al., 2006) and is considered

pharmacologically active as well (Borges et al., 2007).

The indirect photolysis of MESO should be considered as well, and it

could be based on the self-sensitization process. Self-sensitization of

phenothiazine derivatives has been pointed out in the study of mono-

chromatic photolysis of N-methyl phenothiazine (Manju et al., 2012).

In general, self-sensitization is concentration dependent, commonly oc-

curring in higher concentrations of the target compound (Werner et al.,

2006). Therefore, the formation of ROS (¹O₂, HO₂•, and H₂O₂) should not

be neglected as proposed by the pathway (xiii) in Fig. 2. Chen et al.

(2008) have investigated the formation of ¹O₂in the photolysis of tetra-

cycline under simulated sunlight conditions, detected ¹O₂over a wide

pH range, with an increase with the of irradiation time. Besides, the for-

mation of H₂O₂in simulated sunlight conditions was also identiﬁed and

proposed to be generated through the O⁻₂• formation (Chen et al., 2008).

On the other hand, Kelly and Arnold (2012) have investigated the self-

sensitization of phytoestrogens and the generation of ¹O₂during pho-

tolysis. They could not detect ¹O₂and argued that such behavior

might be due to lower energy and short-lived triplet states of the parent

compounds. In their study dissolved oxygen acted mainly as a triplet

quencher making direct photolysis the main photolytic pathway

(Kelly and Arnold, 2012). It has also been proposed that self-

sensitization processes are unlikely to happen at environmental con-

centrations of the parent compounds (Wammer et al., 2013). On the

other hand, in the aquatic environment, indirect photolysis occurs due

to the presence of other molecules such as dissolved organic matter in-

cluding humic acids and fulvic acids as well as nitrate or carbonate that

can act as sensitizers under sunlight leading to ROS (Jacobs et al., 2012;

Li et al., 2016; Lin et al., 2013).

Hydrogen abstraction or dehydroxylation have also taken place

forming the TP 385. Due to the absence of the product ion of m/z 126,

the formation of a double bond was proposed to occur on the piperidine

moiety. Two peaks with the same m/z [M + H]⁺of 373 Da were found,

and based on the HRMS two different structures were proposed. TP 373

I (exact m/z [M + H]⁺373.1573 Da) was proposed to present the for-

mula C₂₀H₂₅O₃N₂S (Δmmu error of −0.780) and be formed by losing

the side chain S(O)CH₃and present the formation of sulfone in the S5-

position followed by a hydroxylation on the phenothiazine ring. The

product ion of m/z 260.0378 (C₁₃H₁₀O₃NS) support the proposed struc-

ture by presenting the three oxygens on the phenothiazine ring, where-

as fragmentation pattern considering the MS²product ions of m/z

293.2015 (C₂₀H₂₅N₂), 180.0813 (C₁₃H₁₀N) and MS³product ion of m/z

179.0730 (C₁₃H₉N) indicated the loss of S(O₂). On the other hand, TP

373 II (exact m/z [M + H]⁺373.1402 Da) was proposed to present the

formula C₂₀H₂₅ON₂S₂(Δmmu error of −0.111) and is assumed to be

formed through an N-demethylation on the piperidine moiety: the

MS²and MS³product ion of m/z 112.1122 Da (C₇H₁₄N) indicated the

loss of CH₃in the R1-substituent.

The TP 371 was proposed to be Thioridazine as they have identical

fragmentation patterns (Text S8, Supplementary material). Formation

of Thioridazine evidences occurrence of an abiotic back-transformation

process in the photolysis of metabolites and it might be assumed to take

place in the environment as well. Back-transformation of metabolite to

parent compound through photolysis have also been reported in the

photolysis from the metabolite 4-nitroso-sulfamethoxazole back to sul-

famethoxazole (Bonvin et al., 2013). A further dehydrogenation step of

MESO takes place forming the TP 369. It is not possible to state the exact

position of the double bond in TP 369 based on the MSⁿdata only. The

formation of TP 369 was also reported in the photolysis of Thioridazine

(Wilde et al., 2016). The TP 357 with the exact of m/z [M + H]⁺

357.1632 Da and calculated elemental composition C₂₀H₂₅O₂N₂S

(Δmmu error of 0.065) is proposed to be formed through sulfoxidation

in the S5-position of the tricyclic phenothiazine ring followed by a hy-

droxylation of an aromatic ring. The fragmentation pattern obtained

by HCD provided the product ion of m/z 126.1278 Da (C₈H₁₆N) indicat-

ing that the R1-substituent was intact and the R2-substituent (S(O)CH₃)

was eliminated, whereas the product ion of m/z 232.0430 Da

(C₁₂H₁₀O₂NS) provided the information that sulfoxidation and hydrox-

ylation occurred on the tricyclic phenothiazine ring.

3.2. Identiﬁcation of TPs and proposal of degradation pathway

A non-target approach and data-dependent acquisition (combina-

tion of full scan and product ion spectra) were applied for the identiﬁca-

tion of transformation products. The samples collected at pre-

determined times were analyzed by a full scan from 60 to 500 m/z in

positive ion mode. A resolving power of 30,000 FWHM was used. The

most intense ion of the full scan was isolated and further fragmented

(MS²) and the two most intense product ions of MS²were isolated

and further fragmented (MS³). A suspect screening list with speciﬁc

masses of the most probable TPs was used in the data-dependent acqui-

sition, whereas in the non-target approach the most intense ion was ac-

tivated if no parent masses were found. The total ion chromatograms,

fragmentation patterns (up to MS³) and proposed structures of the

TPs can be seen in Text S7 and Text S8 (Supplementary material).

Two carbazole ring derivatives were found during the photolysis of

MESO, TP 355 and TP 337 (exact m/z [M + H]⁺355.1841 Da and [M

+ H]⁺337.1727 Da, elemental composition, calculated formula

Please cite this article as: Wilde, M.L., et al., Combination of experimental and in silico methods for the assessment of the phototransformation

products of the antipsychotic drug/metabolite Mesoridazine, Sci Total Environ (2017), https://doi.org/10.1016/j.scitotenv.2017.08.040

M.L. Wilde et al. / Science of the Total Environment xxx (2017) xxx–xxx

Fig. 3. Proposed photolysis pathway of MESO under Xe lamp irradiation. * means the chiral centers.

C₂₁H₂₇ON₂S (Δmmu error of 0.249) and C₂₁H₂₅N₂S (Δmmu error of

−0.626), respectively). In accordance with their fragmentation pat-

terns, a sulfoxide elimination on the S5-position can be assumed for

both TPs. Carbazole derived TPs were also found in the photolysis of Thi-

oridazine (Wilde et al., 2016) through sulfoxide elimination. The forma-

tion of a carbazole ring during photolysis was reported for the

photolysis of N-methyl phenothiazine (Manju et al., 2012) and in the

photolysis of diclofenac (Musa and Eriksson, 2009). Besides,

carbazole-derived TPs were also identiﬁed in a previous work regarding

the photolysis of Thioridazine in the same conditions (Wilde et al.,

2016), indicating that both parent compound (Thioridazine) and its

human metabolite (Mesoridazine) can lead to carbazole-derivative

compounds by long-term exposition to irradiation.

With exception of TP 373 I, TP 357, TP 355, and TP 337 the photolysis

of MESO have not produced many changes in the pharmacophore struc-

ture of the phenothiazine pharmaceuticals. Phenothiazine pharmaceu-

ticals are composed of the phenothiazine tricyclic ring (represented in

blue), an alkyl substituent on N10-position (represented in black), and

an electron withdrawing group bonded to C2-position (represented in

red) (Jaszczyszyn et al., 2012). However, the ring substitutions interfere

in the antipsychotic activity, being that the substitution on C1- and C4-

position interfere in the bending of the side chain and the S-binding af-

ﬁnity towards the receptor, respectively (Alagarsamy, 2010; Sriram and

Yogeeswari, 2010). More than one substitution on the phenothiazine

ring decreases the antipsychotic potency (Alagarsamy, 2010; Sriram

and Yogeeswari, 2010), i.e. although speculative, TP 403 VI, VIII might

be less potent than MESO and Thioridazine. The oxidation on the S5-

position to sulfoxide and sulfone decreases the activity (Wójcikowski

et al., 2006) and, consequently, TP 357 and TP 373 I can be expected

as of less potency. Besides, the hydroxyl group is also a strong

electron-donating group contributing to reduce the activity.

The alkyl side chain modulates the pharmacological antipsychotic

activity (Faria et al., 2015). The substitution on the α-C tends to de-

crease the antipsychotic potency (Alagarsamy, 2010; Sriram and

Yogeeswari, 2010). Thus, it is expected that TP 403 I presents lower po-

tency that MESO. Depending on the substituent in the β-C, it might in-

crease or decrease the activity (Alagarsamy, 2010; Sriram and

Yogeeswari, 2010). Therefore, TP 403 V might also have activity. Pheno-

thiazines with the tertiary amino group have high potency (Jaszczyszyn

et al., 2012) and, consequently, TP 373 II might present lower potency

than MESO. Likewise, TP 403 II-IV, VII that was proposed to undergo hy-

droxylation on the piperidine moiety and TP 385 and TP 369 with a pro-

posed double bond in the piperidine moiety could also present higher or

lower activity than MESO and Thioridazine, which would need further

tests to evaluate it.

According to the TPs proposed, the photolysis of MESO leads to new

chiral compounds, which can possess different pharmacological activi-

ties and toxicities (Kasprzyk-Hordern, 2010). Mesoridazine has two chi-

ral centers and the photolysis lead to both the loss of one chiral center

and to the formation of TPs with new chiral centers. The formation of

TPs 403 I\\V and VII present a new chiral center in the piperidine moi-

ety. This third chiral point occurs in the position that the hydroxylation

takes place at α- or β-carbon. The formation of TPs 403 VI and VIII, TP

385, TP 373 II, TP 357 and TP 355 maintained the same number of chiral

center as the parent compound. On the other hand, the formation of TP

403 X, TP 373 I, TP 371 (Thioridazine), TP 369, and TP 337 present a loss

Please cite this article as: Wilde, M.L., et al., Combination of experimental and in silico methods for the assessment of the phototransformation

products of the antipsychotic drug/metabolite Mesoridazine, Sci Total Environ (2017), https://doi.org/10.1016/j.scitotenv.2017.08.040

M.L. Wilde et al. / Science of the Total Environment xxx (2017) xxx–xxx

of one chiral center. Besides, TP 385, TP 369, and TP 357 depending on

the position of the double bond in the piperidine moiety might lose

the chiral center.

and the stereochemistry of the sulfoxide moiety was shown to play an

important role in the structure-activity relationship (Choi et al., 2004).

Fig. 4 shows the normalized chromatographic peak area ratio A/A₀

(%), where A is the relative peak area of the TP and A₀is the relative

peak area of MESO before photolysis. Although it is not possible to relate

the peak area ratio with a concentration in the case of TPs since no stan-

dards are available, it is possible to relate the peak area ratio with the

formation and further transformation of TPs during the photolysis.

Thus, TP 403 III, IV and V have achieved up to 15.6%, 8.4%, and 19.8% of

the initial area of the parent compound, respectively. As observed,

most of the TPs presented a tendency in increase the peak area with

the evolution of photolysis.

Thioridazine is used as a racemate, but the (R)-enantiomer has been

shown to be 2.7 times more potent than the (S)-enantiomer of binding to

the D2 dopamine receptor, and nearly ﬁve times more potent than

alpha-1 receptor antagonist (Choi et al., 2004; Leonard, 2003). On the

other hand, the S-isomer has a 10-fold greater afﬁnity for the D1 receptor

than the R-form. Besides, dose-response relationship showed that the ra-

cemate is 12 times more potent than S-isomer and 3 times more potent

than the R-isomer (Leonard, 2003). The enantiomers of MESO have

shown different binding afﬁnities towards different receptors as well,

25 50 75 100 125 150 175 200 225 250 275

Time (min)

25 50 75 100 125 150 175 200 225 250 275

25 50 75 100 125 150 175 200 225 250 275

Time (min)

25 50 75 100 125 150 175 200 225 250 275

25 50 75 100 125 150 175 200 225 250 275

Time (min)

25 50 75 100 125 150 175 200 225 250 275

25 50 75 100 125 150 175 200 225 250 275

Time (min)

25 50 75 100 125 150 175 200 225 250 275

Time (min)

Fig. 4. Proﬁle of the normalized peak area with A/A₀(%) of the Transformation Products (TPs) during the photolysis by means of Xe lamp (TXE 150 W) irradiation. Data obtained by using

the extracted ion chromatogram acquired through LC-HRMS in full scan mode (50–500 m/z). Initial conditions: [MESO·besylate] 50 mg L⁻¹(35.48 mg L⁻¹of MESO only), pH 7.0, temp.:

20 2 °C (n = 2).

Please cite this article as: Wilde, M.L., et al., Combination of experimental and in silico methods for the assessment of the phototransformation

products of the antipsychotic drug/metabolite Mesoridazine, Sci Total Environ (2017), https://doi.org/10.1016/j.scitotenv.2017.08.040

M.L. Wilde et al. / Science of the Total Environment xxx (2017) xxx–xxx

The A/A₀proﬁle of TPs 403 III and V have shown an increase up to

giving evidence that the biodegradation found in CBT (in terms of

BOD) can be attributed to the counterion besylate.

150 min of photolysis followed by a further transformation. The peak

area proﬁle of TPs has indicated that the preferential degradation path-

way in the beginning of photolysis was through hydroxylation/

sulfoxidation, while other TPs were formed later as can be seen in Fig.

4. The TP 371, TP 360, and TP 355 were detected after 16 min of photol-

ysis only, whereas TP 385, TP 373 I and II, and TP 357 were noticed after

64 min of photolysis, and TP 337 only appeared after 150 min of

photolysis.

Regarding the EIC peak area results of MRT, which demands a higher

inoculum volume, a different behavior was observed for MESO (Fig. 5

(C)). As observed, TPs were again present in this sample, a decrease in

peak area of MESO was followed by an increase in the peak area of TPs

403 II and III. The same behavior was also observed in the toxicity con-

trol (Text S9, Supplementary material). However, in the sterile control,

the EIC peak area results were similar as those found in CBT, MESO pre-

sented an increase in peak area, which was followed by a decrease in the

peak area of the TPs in this sample (Text S9, Supplementary material).

Fig. 5 (D) depicts the results of the sample taken after 256 min of

photolysis and submitted to MRT. The TP 403 V underwent a reduction

in peak area after 28 days of the test. No signiﬁcant changes in peak area

were observed for MESO and most of the TPs present this sample were

stable during the test. The sterile control showed a reduction in chro-

matographic peak area for the most of TPs indicating that abiotic pro-

cesses as described above take place.

MESO has a tricyclic ring and an electron-withdrawing moiety

(S(O)CH₃) as the R2-substituent group and a hydrophilic side chain as

an R1-substituent group. According to the ‘rules of thumb’ for biodegra-

dation (Boethling et al., 2007) such molecular features do not favor bio-

degradability. The introduction of an oxygen atom can as hydroxyl

group favor aerobic biodegradation (Boethling et al., 2007), which was

observed other studies (Rastogi et al., 2015, 2014). The photolysis of

MESO led to TPs presenting hydroxylation and sulfoxidation, but bio-

degradation was not observed in the present study. However, abiotic

processes were the main responsible for the reduction in chromato-

graphic peak area, which is in agreement with the biodegradation

tests applied to phenothiazine-derived Pharmaceuticals (Trautwein

and Kümmerer, 2012a, 2012b; Wilde et al., 2017, 2016).

3.3. Aerobic biodegradability of phototransformation products

The validity criteria of CBT and MRT were fulﬁlled (as for data see

Text S9, Supplementary material). Biodegradation of 23.3% and 18.6%

was observed in CBT regarding O₂consumption (i.e. biochemical oxygen

demand, BOD) for MESO⋅besylate at 0 min and after 256 min of photol-

ysis, respectively (Table 1). Given the natural variation in the biological

CBT test system that is no difference. According to the OECD 301 test se-

ries, a substance is considered readily biodegradable if N60% of ThOD is

achieved after 28 days of the test (OECD, 1992). Consequently, MESO⋅

besylate has to be classiﬁed as not readily biodegradable. In a CBT,

which could also be called a BOD₂₈test (biological oxygen demand

after 28 days), performed in our laboratory earlier, biodegradation of

pure benzenesulfonic acid (besylate) was 25.0% ( 5.65%) after

28 days (Fig. S3, Supplementary material). Benzenesulfonic acid

showed to be resistant to BOD₅(20 °C) biodegradation, however bacte-

ria present in raw municipal sewage degraded benzenesulfonic acid

after an adaptation period (Janeczko and Gomólka, 1990). Besides,

benzenesulfonic acid was found to be biodegradable in the Japanese

MITI test (Kawasaki, 1980). Therefore, it can be assumed that the ob-

served biodegradation of MESO⋅besylate is mainly due to the biodegra-

dation of the besylate counterion itself and MESO is not biodegradable

at all.

3.4. Bacterial toxicity of MESO and its photolytic TPs

In the MRT Mesoridazine besylate achieved 6.4% and 0.7% of biodeg-

radation at 0 min and after 256 min of photolysis, respectively (Table 1).

That is no signiﬁcant difference and relates to “not biodegradable”. The

toxicity control vessels in both tests indicated no toxicity according to

the OECD guidelines since N25% of biodegradation was reached in the

controls (OECD, 1992).

In order to assess if MESO and TPs undergo any transformation dur-

ing the biodegradation tests, samples collected during the tests were an-

alyzed by LC-ESI-IT-MSⁿ. The photo-instability, presence of dissolved

oxygen, and autocatalytic characteristics of phenothiazine-derived

Pharmaceuticals (Manju et al., 2012; Nałecz-Jawecki et al., 2008) led

some TPs of MESO were already present in the sample before photolysis

(0 min, Fig. 5 A and 5C).

Fig. 5 (A) shows the peak area results of the CBT. It was found that

the peak area of MESO did not decrease throughout the test. Instead, it

increased. Back-transformation of TPs to MESO could be an explanation

for this since the increase in peak area of MESO goes along with a de-

crease in peak area of the TPs present in the same sample. The CBT re-

sults of the sample collected after 256 min of photolysis and

containing at most TPs can be seen in Fig. 5 (B). No signiﬁcant differ-

ences were observed in terms of peak area for the TPs after 28 days of

CBT (i.e. not biodegradable). MESO was stable in the complex mixture,

The concentration-response relationships of MESO in the modiﬁed

LBT and the parameters of the ﬁtted curves are presented in Text S10

(Supplementary material). The resulting EC₅₀values were 18.1

μmol L⁻¹(30 min) and 25.4 μmol L⁻¹(24 h) for luminescence inhibi-

tion and 211.2 μmol L⁻¹for growth inhibition. The measured concen-

tration of MESO and the A/A₀proﬁle of the TPs in the reaction mixture

used for toxicity testing is presented in Fig. 1 (A) (50.00 mg L⁻¹

MESO, i.e. 70.46 mg L⁻¹of MESO⋅besylate) and Text S11 (Supplemen-

tary material), respectively.

The irradiation of MESO resulted in a moderate reduction of

short-term and long-term luminescence inhibition (Figs. 6 (A) and

6 (B)). In both cases, the observed effect of the photolysis mixture

was well explained by the predicted effect of MESO over the whole

treatment process. In the case of growth inhibition, it was not possi-

ble to identify any activity changes during the treatment process,

since the observed effect was below the 20% threshold for signiﬁ-

cance in all tested samples (Fig. 6 (C)). In conclusion, the observed

residual effect of the photolytic mixture was most probably caused

by MESO alone, which is in agreement with the ﬁnding that the mix-

ture of TPs as a whole possessed a considerably lower bacterial tox-

icity than the parent compound.

Table 1

Results of the investigated aerobic biodegradation test assays applied for Mesoridazine besylate before and after 256 min of photolysis by means of Xe lamp (TXE 150 W) irradiation.

Biodegradation test

Substance

Test sample

(min)

Biodegradation after

28 days (%)

Biodegradation in toxicity

control after 28 days (%)

Degradation in the sterile

control after 28 days (%)

DOC elimination

(%)

Closed bottle test

(OECD 301 D)

Manometric respirometry test

(OECD 301 F)

Mesoridazine besylate

256

23.25 ( 1.20)

18.60 ( 3.20)

6.39 ( 5.42)

0.72 ( 2.59)

48.10 ( 0.62)

45.57

43.94

–

2.67

1.00

–

16.9

20.4

256

34.94

Please cite this article as: Wilde, M.L., et al., Combination of experimental and in silico methods for the assessment of the phototransformation

products of the antipsychotic drug/metabolite Mesoridazine, Sci Total Environ (2017), https://doi.org/10.1016/j.scitotenv.2017.08.040

M.L. Wilde et al. / Science of the Total Environment xxx (2017) xxx–xxx

Transformation products

Transformation Products

Fig. 5. LC-MS/MS peak areas of Mesoridazine (MESO) and Transformation Products (TPs) before and after 28 days of aerobic biodegradation. (A) Closed Bottle Test of MESO before

photolysis (i.e. t = 0 min); (B) Closed Bottle Test of MESO and TPs after 256 min of photolysis; (C) Manometric Respirometry Test of MESO before photolysis (i.e. t = 0 min); and

(D) Manometric Respirometry Test of MESO and TPs after 256 min of photolysis (n = 2).

3.5. Screening of potential PBT/vPvB compounds among the TPs of

Mesoridazine

Text S15 (Supplementary material). Fig. 7 presents the results by plot-

ting the US-EPA PBT proﬁler index versus the QSARINS Insubria-PBT

index. The QSARINS Insubria-PBT index predictions have been reported

as more precautionary than US-EPA PBT index (Gramatica et al., 2015).

The TPs are divided into three groups by reference lines (18 for US-EPA

PBT proﬁler and 1.5 for the QSARINS Insubria-PBT index). The group

(I) can be classiﬁed as non-PBT compounds, once both separated in silico

QSAR indexes have classiﬁed them in this order. The TPs classiﬁed in

this group were the most of the stereochemical structures of the TPs

403 (I-IX) and TP 373 I, either being the result of hydroxylation or

sulfoxidation MESO. TP 403 IX was proposed to be Sulforidazine, a

human metabolite of MESO formed in phase I of the metabolic process

and as described above is pharmacologically active (Borges et al.,

2007). As expected, the sulfoxidation and/or hydroxylation produce

an increase in molecule polarity, consequently, decrease the log K_OW

and log BCF values. Consequently, such TPs are more hydrophilic and

would be dispersed in the aquatic environment with no tendency to

bioaccumulation.

Regarding compounds with known chemical structures but un-

known properties and effects, the use of in silico QSAR predictions is a

valuable tool in their risk assessment. Gramatica et al. (2015) have pro-

posed a consensus among the in silico QSAR predictions of US-EPA PBT

proﬁler and the QSARINS Insubria-PBT index regarding the screening

of PBT/vPvB compounds. The criteria adopted in both US-EPA PBT pro-

ﬁler and QSARINS Insubria-PBT index can be seen in Text S13 and Text

S14 (Supplementary material). With that in mind, we have applied

such screening methodology to the TPs formed from the photolysis of

MESO in order to identify possible PBT/vPvB compounds among the TPs.

All structures presented values below the QSARINS cutoff value

(0.0833) and were in the applicability domain (AD) of the model. The

exception was the TP 403 VIII alt, which presented a value higher than

the cutoff (0.0928), not being within the AD. Accordingly, the different

possible structures of the TPs were submitted to both PBT proﬁler and

QSARINS software using the same classiﬁcation methodology proposed

by Gramatica and co-workers (Cassani and Gramatica, 2015; Gramatica

et al., 2015; Sangion and Gramatica, 2016). The in silico predictions of

US-EPA PBT proﬁler and QSARINS Insubria-PBT index can be seen in

The group (II) shows the TPs which were classiﬁed as PBT com-

pounds by US-EPA PBT proﬁler but not by the in silico predictions of

QSARINS Insubria-PBT index. This could be overestimated PBT com-

pounds according to Gramatica et al. (2015). The parent compound

Please cite this article as: Wilde, M.L., et al., Combination of experimental and in silico methods for the assessment of the phototransformation

products of the antipsychotic drug/metabolite Mesoridazine, Sci Total Environ (2017), https://doi.org/10.1016/j.scitotenv.2017.08.040

M.L. Wilde et al. / Science of the Total Environment xxx (2017) xxx–xxx

Fig. 6. Observed effect values (mean SD) of the photolytic mixture of 129.3 μmol L⁻¹Mesoridazine besylate (MESO) in comparison to the predicted effect of residual MESO by means of

short-term luminescence inhibition after 30 min (A), long-term luminescence inhibition after 24 h (B) and growth inhibition (C) in the modiﬁed luminescent bacteria test. The positive

control samples contained 9 mg L⁻¹of 3,5-Dichlorophenol (3,5-DCP) and 0.2 mg L⁻¹Chloramphenicol (CAM). The ﬁnal concentration in the test medium of the photolytic mixture and

the positive control samples was 50% (v/v).

MESO and TPs in this group resulted from dehydroxylation (TP 371, Thi-

oridazine), dehydrogenation (TP 385), both dehydroxylation and dehy-

drogenation (TP 369), demethylation (TP 373 II) and even the

hydroxylated variants from TP 403 (VI, VIII) with OH attachment at the

C1- and C9-position of the phenothiazine moiety. The predictions of

A/A₀ratio and after 16 min and 150 min of photolysis, respectively.

The proﬁle of A/A₀(see Fig. 4) indicates that these TPs show a tendency

to increase with longer exposition times, which is in agreement with the

observation of Manju et al. (2012). Moreover, carbazole-derived TPs

have also been reported as TPs from the degradation of diclofenac

(Musa and Eriksson, 2009; Salgado et al., 2013). Although not the

same compounds as the carbazole derivative TPs found in this study, ha-

logenated derived carbazoles have shown persistence and dioxin-like

toxicity in soil (Mumbo et al., 2014), providing an indication of the

PBT potential of carbazole derivatives. Therefore, a more prominent as-

sessment of the carbazole derivative TPs should be further considered.

PBT proﬁler for TP 403 VI and VIII provided higher values for log K_OW

log BCF and lower values for toxicity (Fish Ch.V.) regarding the hydrox-

ylation on C1- and C9-position of the phenothiazine ring. On the other

hand, QSARINS Insubria-PBT index values for these TPs indicated no con-

siderable difference among the possible different positions where the

hydroxylation could take place on the phenothiazine tricycle ring.

The group (III) in Fig. 7 contain TP 337 and TP 355 being PBT com-

pounds according to both indexes and can be considered as PBT/vPvB

compounds in a consensus mode (i.e. both indexes (QSARINS Insubria-

PBT index and US-EPA PBT index agreed in the predictions). These two

TPs are carbazole derivatives. TP 355 and TP 337 were formed in lower

3.6. QSAR predictions of genotoxicity and mutagenicity

The in silico QSAR predictions considering all structures of the pro-

posed TPs including possible isomers are given in Table 2. SMILES of

Please cite this article as: Wilde, M.L., et al., Combination of experimental and in silico methods for the assessment of the phototransformation

products of the antipsychotic drug/metabolite Mesoridazine, Sci Total Environ (2017), https://doi.org/10.1016/j.scitotenv.2017.08.040

M.L. Wilde et al. / Science of the Total Environment xxx (2017) xxx–xxx

Table 2

In silico QSAR predictions for mutagenicity of Mesoridazine and all possible different struc-

tures proposed for the TPs formed during photolysis according to QSAR models provided

by Multicase CASE Ultra™ and Leadscope Model Applier™ (A-J: Software models used, as

for details see at the end of the table).

Structure ID^a

QSAR predictions

Case Ultra^b

Leadscope^c

Mesoridazine

TP 403 I

−

TP 403 VI, VIII b-f

TP 403 II, III, IV, VII a-d

TP 403 V

TP 403 II, III, IV, VII a

TP 403 II, III, IV, VII b

TP 403 II, III, IV, VII c

TP 403 II, III, IV, VII d

TP 403 V

TP 403 VI, VIII a

TP 403 VI, VIII b

TP 403 VI, VIII c

TP 403 VI, VIII d

TP 403 VI, VIII e

TP 403 VI, VIII f

TP 403 VI, VIII g

TP 403 IX

QSARINS Insubria-PBT index

OD OD

Fig. 7. The plot of US EPA PBT proﬁler index versus QSARIN Insubria-PBT index for the

identiﬁcation of potential PBT/vPvB compounds among the TPs formed from MESO by

photolysis.

−

TP 403 VIII alt

TP 373 I a

TP 373 I b

TP 373 I c

TP 373 I d

OD OD

−

OD OD OD OD

OD OD OD

OD OD OD OD

all structures proposed can be seen in Text S12 (Supplementary materi-

al). MESO and most of the TPs formed have been classiﬁed as “inconclu-

sive”, “out of domain” and “negative” alerts. The “inconclusive” alert

means that the calculated probability of the proposed structure being

positive falls inside the so-called “gray zone”, which depends on the

model applied, whereas “out of domain” means that the structure tested

is not covered by the applicability domain of the applied QSAR model.

The “negative” alerts point out that that the analyzed structure has a cal-

culated probability of being positive lower than the model's classiﬁca-

tion threshold (50.0%) and is not within the “gray zone”. In other

words, predicted negative alerts indicate that the compounds are classi-

ﬁed as not presenting genotoxicity and mutagenic activities according

to their molecular structure. On the other hand, “positive” alerts indi-

cate that the probability of the tested structure being positive as for

the endpoint tested is higher than the model's classiﬁcation threshold

(50.0%) and is not within the “gray zone”.

The “Konsolidator” (column F in Table 2) is a tool of the CASE Ultra

software intended to generate evidence for review and regulatory issues

by combining and re-evaluating the multiple statistical and expert rule

models. The “Konsolidator” considers the following features: (i) It is

based on the multiple statistical and expert rule models; (ii) It compares

the tested compounds with active pharmaceuticals ingredients (API) or

inert compounds; (iii) Comprehensive literature reference in support of

the data is provided; (iv) Automatic neighborhood analysis of the tested

compound is performed; (v) Evaluates the alerts and their validity; (vi)

Assesses unknown fragments; and (vii) Increase in the coverage of the

in silico predictions by resolving “out of domain” and “inconclusive”

alerts is provided.

−

OD OD OD

−

TP 373 II

TP 371 (Thioridazine) IN

−

TP 385 a

TP 385 b

TP 385 c

TP 385 d

TP 385 e

TP 385 f

TP 369 a

TP 369 b

TP 369 c

TP 369 d

TP 369 e

TP 369 f

TP 357 a

TP 357 b

TP 357 c

TP 357 d

TP 355

−

OD OD

OD OD OD

OD OD OD OD

OD OD OD

−

OD OD

TP 337 a

TP 337 b

TP 337 c

TP 337 d

TP 337 e

TP 337 f

OD OD OD OD

OD (Out of Domain): tested chemical is not covered by the applicability domain of the

model.

IN (Inconclusive): the calculated probability of being positive falls inside the “gray zone”

(35%–65% of probability for A and 40%–60% of probability for B, C, D, and E).

(+) positive alert (N60%). (−) negative alert (b40%).

The SMILES codes of the structure ID of the TPs can be seen in Text S12 (Supplemen-

Based on the “Konsolidator” features, MESO and most of TPs were

classiﬁed as presenting negative predictions for bacterial mutagenicity

as can be observed in Table 2. However, two TPs, namely TP 355 and

TP 337, presented statistically and rule-based positive alerts for mutage-

nicity in bacteria according to the CASE Ultra models applied, indicating

that these TPs might have structural features closely linked to DNA-

reactivity. Positive prediction for bacterial mutagenesis towards Salmo-

nella typhimurium was also pointed out by Leadscope Model Applier

models. Additionally, the Leadscope software has pointed out negative

predictions for MESO and the most of TPs proposed in the Mammalian

mutation model. The same trend was also observed the Leadscope

models regarding the genotoxic activities of the models in vitro chromo-

some aberrations average model and in vivo Micronucleus single model.

TP 355 and TP 337 are carbazole derivatives TPs. Fig. 8 depicts the

formation of the carbazole tricyclic ring according to Manju et al.

(2012), and highlight the structural alerts (in bold) for bacterial

tary material). The letters represent different isomers of TPs with the same m/z (i.e. OH or

double bond position).

Case Ultra models according to ICH guideline M7: (A) GT1_A7B (Salmonella t. 7-strains

(RCA)); (B)

GT1_AT_ECOLI

(E.

coli/Salmonella TA102

(A-T

Base

Pair

Mutation)); (C) GT_EXPERT (Expert Rules for Bacterial Mutagenicity); (D) PHARM_ECOLI

(E.coli mutagenicity (all strains)); (E) PHARM_SALM (Salmonella mutagenicity

(TA97,98,100,1535-1538)); (F) “Konsolidator” Outcome.

Leadscope models: (G) In Vitro Chromosome aberrations average

model; (H) Mammalian mutation; (I) In vivo Micronucleus single model; (J) Bacterial

mutagenesis towards Salmonella typhimurium.

mutagenicity provided by the CASE Ultra models. TP 355 and all possible

different structures proposed for TP 337 have the same structural alerts

with high statistical signiﬁcance, which is because of the shared carba-

zole backbone.

The predicted mutagenic activity of carbazole derivatives is support-

ed by experimental results. Carbazole derivatives might have caused

Please cite this article as: Wilde, M.L., et al., Combination of experimental and in silico methods for the assessment of the phototransformation

products of the antipsychotic drug/metabolite Mesoridazine, Sci Total Environ (2017), https://doi.org/10.1016/j.scitotenv.2017.08.040

M.L. Wilde et al. / Science of the Total Environment xxx (2017) xxx–xxx

R₁

R₂

R₁

¹⁰N

[c.]:n:[c.](:cH):[c.]

R₂

R₁

R₂

O₂

R₁

R₁

R₂

O₂

R₁

R₁

R₂

O₂

R₁

R₁

R₂

R₁

(Salmonella

(TA97,98,100,1535-1538))

R₂

cH:[c.]:[c.](:[c.]):cH:cH

Fig. 8. Formation of a carbazole ring through photolysis (adapted from Manju et al., 2012) and the structural alerts (in bold) responsible for the positive alerts in the carbazole tricyclic ring

of TP 355 and TP 337 provided by CASE Ultra software according to the ICH M7 guideline.

DNA mutation in a male germ cell of Swiss albino mice (Jha and Bharti,

2002), and some methyl-, nitro- and amino-substituted carbazoles

were found to be mutagenic to Salmonella typhimurium (André et al.,

1997, 1995; Grover and Bala, 1993; LaVoie et al., 1982). One of the mo-

lecular descriptors that are correlated to the mutagenic activity of carba-

zole derivatives is the hydrophobicity (André et al., 1997).

kind of TPs as Thioridazine, but in less number and with structural dif-

ferences due to the different precursor compounds. The same kind of

mechanism was veriﬁed for both compounds by means of Xe lamp

(TXE 150 W) irradiation. This also shows that the same TPs can evolve

from the degradation of different compounds. New TPs were found as

well, consequently, this new mixture of TPs was further analyzed in aer-

obic biodegradability tests and as for toxicity. Consequently, an isolated

assessment of related APIs may underestimate the true risk of common

TPs.

3.7. Environmental signiﬁcance and implications

The isolation and/or synthesis of so many TPs to allow separately

tests for an extended assessment is impracticable for ﬁnancial reasons.

Thus, this study aimed to better understand the signiﬁcance of the TPs

formed and present a workﬂow how this can be done. The screening ap-

proach as presented in the paper helps to resolve this issue as it allows

for collecting basic information and the selection of compounds being of

most interest for a PBT/vPvB assessment thereby allowing including TPs

in such an assessment. The use of high initial concentrations enables to

generate a broad range of TPs at levels that would allow an analytical

detection and studying biodegradability as well as toxicity experimen-

tally for proactive assessment of fate and effects. Especially because of

the sunlight simulating photolysis has been shown to be a mere

phototransformation process (Wang and Lin, 2014). Photo TPs are of in-

terest if they are not biodegradable in the environment. The use of envi-

ronmentally relevant concentrations would make these tests unfeasible,

due to the high ThOD required for such tests. However, biodegradability

is an important parameter for environmental risk assessment as well as

for sustainability and green chemistry, once the development of new

pharmaceuticals compounds should focus on their biodegradability,

minimizing the exposure of ecosystems and, consequently, humans to

pharmaceuticals (Kümmerer, 2007; Leder et al., 2015; Rastogi et al.,

2015).

The evidence of photolytic back-transformation from MESO to Thio-

ridazine by means of Xe lamp (TXE 150 W) irradiation in this manu-

script provides evidence that photolysis through natural sunlight

might not entirely attenuate the presence of the parent drugs in the en-

vironment, but to some extent even may regenerate them. This might

be especially important in cases where the metabolite shows a much

lower pharmacological or toxicological activity than its precursor parent

drug. As an example, Thioridazine was determined to be approximately

18-fold more active than MESO regarding the inhibition of bacterial

growth (Wilde et al., 2016). Therefore, research on the contribution of

natural abiotic process on human metabolites and TPs as a possible

back-transformation process should be further investigated.

The high initial concentration of MESO allowed for the experimental

study of the ready biodegradability by applying two OECD standardized

tests, CBT (OECD 301D) and MRT (OECD 301F), which have indicated

that MESO and the mixture of its photo TPs are not biodegradable.

That implies that they persist in the environment as they are stable

end products of photolysis too. The decrease in concentration of MESO

and the formation of hydroxylated and sulfoxide as the main TPs had

led to a decrease in the ecotoxicity towards V. ﬁscheri. However, psychi-

atric drugs might alter the composition of freshwater invertebrate com-

munities, as was the case of carbamazepine (Jarvis et al., 2014a, 2014b).

The pressure for identiﬁcation of possible hazardous compounds

and their substitution for greener (i.e. safer) ones is urgently needed

(Alves et al., 2016). The identiﬁcation of possible hazardous TPs is a

In comparison to the photolysis of Thioridazine (Wilde et al., 2016),

an active pharmaceutical ingredient precursor of MESO in the human

body, the photolysis of the human metabolite MESO produced a similar

Please cite this article as: Wilde, M.L., et al., Combination of experimental and in silico methods for the assessment of the phototransformation

products of the antipsychotic drug/metabolite Mesoridazine, Sci Total Environ (2017), https://doi.org/10.1016/j.scitotenv.2017.08.040

M.L. Wilde et al. / Science of the Total Environment xxx (2017) xxx–xxx

necessity in this context. The combination of experimental work and in

References

silico QSAR models had proved to be a powerful screening approach in

the risk assessment of unknown compounds such as TPs as demonstrat-

ed in this study allows for a better understanding of environmental risks

and even to further prioritize possibly hazardous compounds for further

testing. Concerning the TPs of MESO found in this study, attention

should be given to the formation of carbazole derivatives by photolysis

indicated as possible mutagenic and putative PBT/vPvB according to

QSAR in silico predictions and literature related to other carbazole deriv-

atives that were observed in the photolysis of both compounds, Thiorid-

azine (Wilde et al., 2016) and MESO.

Alagarsamy, V., 2010. Textbook of Medicinal Chemistry. 1st ed. Elsevier India, New Delhi,

India.

Alves, V.M., Muratov, E.N., Capuzzi, S.J., Politi, R., Low, Y., Braga, R.C., Zakharov, A.V.,

Sedykh, A., Mokshyna, E., Farag, S., Andrade, C.H., Kuz'min, V.E., Fourches, D.,

Tropsha, A., 2016. Alarms about structural alerts. Green Chem. 18:4348–4360.

https://doi.org/10.1039/C6GC01492E.

Amaral, L., 2012. Thioridazine: an old neuroleptic effective against totally drug resistant

tuberculosis. Acta Medica Port. 25, 118 –121.

André, V., Boissart, C., Sichel, F., Gauduchon, P., Le Talaër, J.Y., Lancelot, J.C., Robba, M.,

Mercier, C., Chemtob, S., Raoult, E., Tallec, A., 1995. Mutagenicity of nitro- and

amino-substituted carbazoles in Salmonella typhimurium. II. Ortho-aminonitro de-

rivatives of 9H-carbazole. Mutat. Res. 345:11–25. https://doi.org/10.1016/0165-

1218(95)90066-7.

André, V., Boissart, C., Sichel, F., Gauduchon, P., Le Talaër, J.Y., Lancelot, J.C., Mercier, C.,

Chemtob, S., Raoult, E., Tallec, A., 1997. Mutagenicity of nitro- and amino-substituted

carbazoles in Salmonella typhimurium. III. Methylated derivatives of 9H-carbazole.

Mutat. Res. 389:247–260. https://doi.org/10.1016/S1383-5718(96)00155-6.

Boethling, R.S., Sommer, E., DiFiore, D., 2007. Designing small molecules for biodegrad-

ability. Chem. Rev. 107:2207–2227. https://doi.org/10.1021/cr050952t.

Bonvin, F., Omlin, J., Rutler, R., Schweizer, W.B., Alaimo, P.J., Strathmann, T.J., McNeill, K.,

Kohn, T., 2013. Direct photolysis of human metabolites of the antibiotic sulfamethox-

azole: evidence for abiotic back-transformation. Environ. Sci. Technol. 47:6746–6755.

https://doi.org/10.1021/es303777k.

Borges, K.B., Borges, W.D.S., Pupo, M.T., Bonato, P.S., 2007. Endophytic fungi as models for

the stereoselective biotransformation of thioridazine. Appl. Microbiol. Biotechnol. 77:

669–674. https://doi.org/10.1007/s00253-007-1171-x.

Cassani, S., Gramatica, P., 2015. Identiﬁcation of potential PBT behavior of personal care

products by structural approaches. Sustain. Chem. Pharm. 1:19–27. https://doi.org/

10.1016/j.scp.2015.10.002.

Celiz, M.D., Tso, J., Aga, D.S., 2009. Pharmaceutical metabolites in the environment: analyt-

ical challenges and ecological risks. Environ. Toxicol. Chem. 28:2473–2484. https://

doi.org/10.1897/09-173.1.

Chakravarti, S.K., Saiakhov, R.D., Klopman, G., 2012. Optimizing predictive performance of

CASE Ultra expert system models using the applicability domains of individual toxic-

ity alerts. J. Chem. Inf. Model. 52:2609–2618. https://doi.org/10.1021/ci300111r.

Chen, Y., Hu, C., Qu, J., Yang, M., 2008. Photodegradation of tetracycline and formation of

reactive oxygen species in aqueous tetracycline solution under simulated sunlight ir-

radiation. J. Photochem. Photobiol. A Chem. 197:81–87. https://doi.org/10.1016/

j.jphotochem.2007.12.007.

Choi, S., Haggart, D., Toll, L., Cuny, G.D., 2004. Synthesis, receptor binding and functional

studies of mesoridazine stereoisomers. Bioorg. Med. Chem. Lett. 14:4379–4382.

https://doi.org/10.1016/j.bmcl.2004.06.078.

Cwiertny, D.M., Snyder, S.A., Schlenk, D., Kolodziej, E.P., 2014. Environmental designer drugs:

when transformation may not eliminate risk. Environ. Sci. Technol. 48, 11737 –11745.

De Gaitani, C.M., Martinez, A.S., Bonato, P.S., 2004. Degradation and conﬁgurational

changes of thioridazine 2-sulfoxide. J. Pharm. Biomed. Anal. 36:601–607. https://

doi.org/10.1016/j.jpba.2004.07.011.

4. Conclusions

The photolysis of the antipsychotic and drug metabolite MESO

through Xe lamp irradiation led to a series of TPs. Sixteen TPs were pro-

posed according to LC-HRMSⁿand, among them, abiotic back-

transformation from MESO to Thioridazine was observed, indicating

that natural attenuation of metabolites could also contribute to the en-

vironmental exposition of the parent compounds. Besides, the photoly-

sis of MESO led to new chiral TPs. Chiral compounds might possess

different pharmacological activities and toxicities, needing further in-

vestigations. No ready biodegradability of the parent compound was

observed in CBT and MRT neither for the photo-TPs was observed. A re-

duction in toxicity towards V. ﬁscheri during photolysis was mainly re-

lated to MESO's decline in concentration. In silico predictions of

QSARINS and US-EPA PBT proﬁler pointed out TP 355 and TP 337 as

PBT/vPvB compounds in a consensus mode. Likewise, statistical and

rule-based in silico predictions for mutagenicity and genotoxicity indi-

cated positive alerts for bacterial mutagenicity in TP 355 and TP 337.

Therefore, this study strongly indicates a possible hazard associated

with the two carbazoles derivative TPs, calling for further investigations

on such derivatives to conﬁrm their hazardous alerts experimentally.

The combination of experimental and in silico tools allowed studying

MESO and TPs as a starting point to better understand the environmen-

tal fate and effects of metabolic compounds. Besides, future studies

should focus beyond the parent compounds and more attention should

be given to metabolites and their TPs. A screening study and a workﬂow

for the assessment of photoproducts combining experimental and in

silico work was performed allowing to get new insights and include

knowledge about TPs of a pharmaceutical present in the environment

and their PBT relevant properties.

Du, Y., Chen, H., Zhang, Y., Chang, Y., 2014. Photodegradation of gallic acid under UV irra-

diation: insights regarding the pH effect on direct photolysis and the ROS oxidation-

sensitized process of DOM. Chemosphere 99:254–260. https://doi.org/10.1016/

j.chemosphere.2013.10.093.

Eap, C.B., Guentert, T.W., Schãublin-Loidl, M., Stabl, M., Koeb, L., Powell, K., Baumann, P.,

1996. Plasma levels of the enantiomers of thioridazine, thioridazine 2-sulfoxide, thi-

oridazine 2-sulfone, and thioridazine 5-sulfoxide in poor and extensive metabolizers

of dextromethorphan and mephenytoin. Clin. Pharmacol. Ther. 59:322–331. https://

doi.org/10.1016/S0009-9236(96)80010-5.

Edhlund, B.L., Arnold, W.A., McNeill, K., 2006. Aquatic photochemistry of nitrofuran anti-

biotics. Environ. Sci. Technol. 40:5422 –5427. https://doi.org/10.1021/es0606778.

Eiduson, S., Geller, E., 1963. The excretion and metabolism of ³⁵S-labeled thioridazine in

urine, blood, bile, and feces. Biochem. Pharmacol. 12, 1429 –1435.

Faria, P.A., Bettanin, F., Cunha, R.L.O.R., Paredes-Gamero, E.J., Homem-de-Mello, P., Nantes,

I.L., Rodrigues, T., 2015. Cytotoxicity of phenothiazine derivatives associated with mi-

tochondrial dysfunction: a structure-activity investigation. Toxicology 330:44–54.

https://doi.org/10.1016/j.tox.2015.02.004.

Fasnacht, M.P., Blough, N.V., 2002. Aqueous photodegradation of polycyclic aromatic hy-

drocarbons. Environ. Sci. Technol. 36:4364–4369. https://doi.org/10.1021/es025603k.

Fatta-Kassinos, D., Vasquez, M.I., Kümmerer, K., 2011. Transformation products of phar-

maceuticals in surface waters and wastewater formed during photolysis and ad-

Acknowledgments

The authors would like to thank the Brazilian program “Science

without borders” from CNPq/CAPES/CsF (Grant Nr. 2367712012-4)

and the Innovations-Inkubator Lüneburg (Teilmaßnahme 1.4, Graduate

School) for the scholarships granted to Dr. M.L. Wilde and J. Menz, re-

spectively. The authors would like to thank A. Haiss and E. Logunova

for planning the aerobic biodegradation tests and S. Hinz for helping

in the toxicity test. Multicase Inc. and Leadscope Inc. are kindly ac-

knowledged for providing the CASE Ultra and Leadscope QSAR pro-

grams, respectively. Dr. P. Gramatica is acknowledged for providing

the QSARINS software (www.qsar.it).

vanced oxidation processes

— degradation, elucidation of byproducts and

assessment of their biological potency. Chemosphere 85:693–709. https://doi.org/

10.1016/j.chemosphere.2011.06.082.

Friedrich, J., Längin, A., Kümmerer, K., 2013. Comparison of an electrochemical and lumi-

nescence-based oxygen measuring system for use in the biodegradability testing ac-

cording to closed bottle test (OECD 301D). CLEAN 41:251–257. https://doi.org/

10.1002/clen.201100558.

Fura, A., 2006. Role of pharmacologically active metabolites in drug discovery and develop-

ment. Drug Discov. Today 11:133–142. https://doi.org/10.1016/S1359-6446(05)03681-

Gramatica, P., Chirico, N., Papa, E., Cassani, S., Kovarich, S., 2013. QSARINS: a new software

for the development, analysis, and validation of QSAR MLR models. J. Comput. Chem.

34:2121–2132. https://doi.org/10.1002/jcc.23361.

Conﬂicts of interest

The authors declared to have no conﬂict of interest.

Appendix A. Supplementary data

Gramatica, P., Cassani, S., Chirico, N., 2014. QSARINS-chem: Insubria datasets and new

QSAR/QSPR models for environmental pollutants in QSARINS. J. Comput. Chem. 35:

1036–1044. https://doi.org/10.1002/jcc.23576.

Supplementary data to this article can be found online at https://doi.

org/10.1016/j.scitotenv.2017.08.040.

Please cite this article as: Wilde, M.L., et al., Combination of experimental and in silico methods for the assessment of the phototransformation

products of the antipsychotic drug/metabolite Mesoridazine, Sci Total Environ (2017), https://doi.org/10.1016/j.scitotenv.2017.08.040

M.L. Wilde et al. / Science of the Total Environment xxx (2017) xxx–xxx

Gramatica, P., Cassani, S., Sangion, A., 2015. PBT assessment and prioritization by PBT

index and consensus modeling: comparison of screening results from structural

models. Environ. Int. 77:25–34. https://doi.org/10.1016/j.envint.2014.12.012.

Grover, I.S., Bala, S., 1993. Mutagenicity of nitro- and amino-substituted carbazoles in Sal-

monella typhimurium. I. Monosubstituted derivatives of 9H-carbazole. Mutat. Res.

299:63–73. https://doi.org/10.1016/0165-1218(93)90120-3.

Hawesx, M., 1993. The metabolism of piperidine-type phenothiazine antipsychotic

agents. III. Mesoridazine in dog, human and rat. Xenobiotica 23. https://doi.org/

10.3109/00498259309059360.

Herrmann, M., Menz, J., Olsson, O., Kümmerer, K., 2015. Identi ﬁ cation of phototransformation

products of the antiepileptic drug gabapentin: biodegradability and initial assessment of

toxicity. Water Res. 85, 11 –21.

International Conference on Harmonization (ICH), 2014. ICH guideline M7. http://

www.ich.org/ﬁ leadmin/Public_Web_Site/ICH_Products/Guidelines/Multidisciplin-

ary/M7/M7_Step_4.pdf, Accessed date: 11 February 2015.

Mumbo, J., Henkelmann, B., Abdelaziz, A., Pﬁster, G., Nguyen, N., Schroll, R., Munch, J.C.,

Schramm, K.W., 2014. Persistence and dioxin-like toxicity of carbazole and

chlorocarbazoles in soil. Environ. Sci. Pollut. Res. 22:1344–1356. https://doi.org/

10.1007/s11356-014-3386-6.

Musa, K.A.K., Eriksson, L.A., 2009. Photodegradation mechanism of the common non-

steroid anti-inﬂammatory drug diclofenac and its carbazole photoproduct. Phys.

Chem. Chem. Phys. 11:4601–4610. https://doi.org/10.1039/b900144a.

Nałecz-Jawecki, G., Hajnas, A., Sawicki, J., 2008. Photodegradation and phototoxicity of

thioridazine and chlorpromazine evaluated with chemical analysis and aquatic or-

ganisms. Ecotoxicology 17:13–20. https://doi.org/10.1007/s10646-007-0171-z.

Obach, R.S., 2013. Pharmacologically active drug metabolites: impact on drug discovery

and pharmacotherapy. Pharmacol. Rev. 65:578–640. https://doi.org/10.1124/

pr.111.005439.

OECD, 1992. Organisation for Economic Co-operation and Development, 2006. OECD

Guideline for Testing of Chemicals 301: Ready Biodegradability. OECD Publishing,

Paris.

Oppenländer, T., 2003. Photochemical Puri ﬁ cation of Water and Air. WILEY-VCH Verlag

GmbH & Co. KGaA, Weinheim, Germany.

Jacobs, L.E., Weavers, L.K., Houtz, E.F., Chin, Y.-P., 2012. Photosensitized degradation of caf-

feine: role of fulvic acids and nitrate. Chemosphere 86:124–129. https://doi.org/

10.1016/j.chemosphere.2011.09.052.

Janeczko, A., Gomólka, E., 1990. Respirometric investigations on the biodegradability of

phenylsulphonic acid. Acta Hydrochim. Hydrobiol. 18:581–590. https://doi.org/

10.1002/aheh.19900180511.

Osorio, V., Imbert-Bouchard, M., Zonja, B., Abad, J.L., Pérez, S., Barceló, D., 2014. Simulta-

neous determination of diclofenac, its human metabolites and microbial nitration/

nitrosation transformation products in wastewaters by liquid chromatography/

quadrupole-linear ion trap mass spectrometry. J. Chromatogr. A 1347:63–71.

https://doi.org/10.1016/j.chroma.2014.04.058.

Paltiel, O., Fedorova, G., Tadmor, G., Kleinstern, G., Maor, Y., Chefetz, B., 2016. Human ex-

posure to wastewater-derived pharmaceuticals in fresh produce: a randomized con-

trolled trial focusing on carbamazepine. Environ. Sci. Technol. 50:4476–4482. https://

doi.org/10.1021/acs.est.5b06256.

Jarvis, A.L., Bernot, M.J., Bernot, R.J., 2014a. Relationships between the psychiatric drug

carbamazepine and freshwater macroinvertebrate community structure. Sci. Total

Environ. 496:499–509. https://doi.org/10.1016/j.scitotenv.2014.07.086.

Jarvis, A.L., Bernot, M.J., Bernot, R.J., 2014b. The effects of the psychiatric drug carbamaze-

pine on freshwater invertebrate communities and ecosystem dynamics. Sci. Total En-

viron. 496:499–509. https://doi.org/10.1016/j.scitotenv.2014.07.086.

Jaszczyszyn, A., Gasiorowski, K., SwiaTek, P., Malinka, W., Cieslik-Boczula, K., Petrus, J.,

Czarnik-Matusewicz, B., 2012. Chemical structure of phenothiazines and their biological

activity. Pharmacol. Rep. 64:16–23. https://doi.org/10.1016/S1734-1140(12)70726-0.

Jha, A.M., Bharti, M.K., 2002. Mutagenic proﬁles of carbazole in the male germ cells of Swiss

albino mice. Mutat. Res. 500:97–101. https://doi.org/10.1016/S0027-5107(01)00303-7.

Jin, H.-E., Lee, K.-R., Kang, I.-H., Chung, S.-J., Shim, C.-K., 2014. Determination of

mesoridazine by liquid chromatography–tandem mass spectrometry and its applica-

tion to pharmacokinetic study in rats. J. Chromatogr. B 958:117–123. https://doi.org/

10.1016/j.jpba.2010.11.001.

Kasprzyk-Hordern, B., 2010. Pharmacologically active compounds in the environment

and their chirality. Chem. Soc. Rev. 39:4466–4503. https://doi.org/10.1039/c000408c.

Kawasaki, M., 1980. Experiences with the test scheme under the chemical control law of

Japan: an approach to structure-activity correlations. Ecotoxicol. Environ. Saf. 4:

444–454. https://doi.org/10.1016/0147-6513(80)90046-9.

Kelly, M.M., Arnold, W.A., 2012. Direct and indirect photolysis of the phytoestrogens ge-

nistein and daidzein. Environ. Sci. Technol. 46:5396–5403. https://doi.org/10.1021/

es300041f.

Papa, E., Gramatica, P., 2010. QSPR as a support for the EU REACH regulation and rational

design of environmentally safer chemicals: PBT identiﬁcation from molecular struc-

ture. Green Chem. 12:836. https://doi.org/10.1039/b923843c.

Rastogi, T., Leder, C., Kümmerer, K., 2014. Designing green derivatives of β-blocker meto-

prolol: a tiered approach for green and sustainable pharmacy and chemistry.

Chemosphere 111:493–499. https://doi.org/10.1016/j.chemosphere.2014.03.119.

Rastogi, T., Leder, C., Kümmerer, K., 2015. Re-designing of existing pharmaceuticals for en-

vironmental biodegradability: a tiered approach with β-blocker propranolol as an ex-

ample. Environ. Sci. Technol. 11756–11763. https://doi.org/10.1021/acs.est.5b03051.

Ravyn, D., Ravyn, V., Lowney, R., Nasrallah, H.a., 2013. CYP450 pharmacogenetic treat-

ment strategies for antipsychotics: a review of the evidence. Schizophr. Res. 149:

1–14. https://doi.org/10.1016/j.schres.2013.06.035.

Roberts, G., Myatt, G.J., Johnson, W.P., Cross, K.P., Blower, P.E., 2000. LeadScope: software

for exploring large sets of screening data. J. Chem. Inf. Comput. Sci. 40:1302–1314.

https://doi.org/10.1021/ci0000631.

Rodrigues, T., Dos Santos, C.G., Riposati, A., Barbosa, L.R.S., Di Mascio, P., Itri, R., Baptista,

M.S., Nascimento, O.R., Nantes, I.I., 2006. Photochemically generated stable cation

radical of phenothiazine aggregates in mildly acid buffered solutions. J. Phys. Chem.

B 110:12257–12265. https://doi.org/10.1021/jp0605404.

Kümmerer, K., 2007. Sustainable from the very beginning: rational design of molecules by

life cycle engineering as an important approach for green pharmacy and green chem-

istry. Green Chem. 9:899. https://doi.org/10.1039/b618298b.

Kümmerer, K., 2009. The presence of pharmaceuticals in the environment due to human

use — present knowledge and future challenges. J. Environ. Manag. 90:2354–2366.

https://doi.org/10.1016/j.jenvman.2009.01.023.

Saiakhov, R., Chakravarti, S., Klopman, G., 2013. Effectiveness of CASE ultra expert system

in evaluating adverse effects of drugs. Mol. Inf. 32:87–97. https://doi.org/10.1002/

minf.201200081.

Saiakhov, R., Chakravarti, S., Sedykh, A., 2014. An improved workﬂow to perform in silico

mutagenicity assessment of impurities as per ICH M7 guideline. Toxicol. Lett. 229:

S164. https://doi.org/10.1016/j.toxlet.2014.06.563.

Kümmerer, K., 2010. Pharmaceuticals in the environment. Annu. Rev. Environ. Resour. 35:

57–75. https://doi.org/10.1146/annurev-environ-052809-161223.

Langford, K., Thomas, K.V., 2011. Input of selected human pharmaceutical metabolites

into the Norwegian aquatic environment. J. Environ. Monit. 13:416–421. https://

doi.org/10.1039/c0em00342e.

LaVoie, E.J., Briggs, G., Bedenko, V., Hoffmann, D., 1982. Mutagenicity of substituted carba-

zoles in Salmonella typhimurium. Mutat. Res. Toxicol. 101:141–150. https://doi.org/

10.1016/0165-1218(82)90004-0.

Leder, C., Rastogi, T., Kümmerer, K., 2015. Putting benign by design into practice-novel

concepts for green and sustainable pharmacy: designing green drug derivatives by

non-targeted synthesis and screening for biodegradability. Sustain. Chem. Pharm. 2:

1–6. https://doi.org/10.1016/j.scp.2015.07.001.

Salgado, R., Pereira, V.J., Carvalho, G., Soeiro, R., Gaffney, V., Almeida, C., Vale Cardoso, V.,

Ferreira, E., Benoliel, M.J., Ternes, T.A., Oehmen, A., Reis, M.A.M., Noronha, J.P., 2013.

Photodegradation kinetics and transformation products of ketoprofen, diclofenac

and atenolol in pure water and treated wastewater. J. Hazard. Mater. 244–245:

516–527. https://doi.org/10.1016/j.jhazmat.2012.10.039.

Sangion, A., Gramatica, P., 2016. PBT assessment and prioritization of contaminants of

emerging concern: pharmaceuticals. Environ. Res. 147:297–306. https://doi.org/

10.1016/j.envres.2016.02.021.

Santos, L.H.M.L.M., Araújo, A.N., Fachini, A., Pena, A., Delerue-matos, C., Montenegro,

M.C.B.S.M., 2010. Ecotoxicological aspects related to the presence of pharmaceuticals

in the aquatic environment. J. Hazard. Mater. 175:45–95. https://doi.org/10.1016/

j.jhazmat.2009.10.100.

Leonard, B.E., 2003. Fundamentals of Psychopharmacology. 3rd Ed. WILEY-VCH, Verlag

GmbH.

Li, Y., Chen, J., Qiao, X., Zhang, H., Zhang, Y.-nan, Zhou, C., 2016. Insights into photolytic

mechanism of sulfapyridine induced by triplet-excited dissolved organic matter.

Chemosphere 147:305–310. https://doi.org/10.1016/j.chemosphere.2015.12.115.

Lin, A.Y.-C., Wang, X.-H., Lee, W.-N., 2013. Phototransformation determines the fate of 5-

ﬂuorouracil and cyclophosphamide in natural surface waters. Environ. Sci. Technol.

47:4104–4112. https://doi.org/10.1021/es304976q.

López-Serna, R., Jurado, A., Vázquez-Suñé, E., Carrera, J., Petrović, M., Barceló, D., 2013. Oc-

currence of 95 pharmaceuticals and transformation products in urban groundwaters

underlying the metropolis of Barcelona, Spain. Environ. Pollut. 174:305–315. https://

doi.org/10.1016/j.envpol.2012.11.022.

Sriram, D., Yogeeswari, P., 2010. Medicinal Chemistry. 2nd ed. Pearson, New Delhi, India.

Thanacoody, H.K.R., 2007. Thioridazine: resurrection as an antimicrobial agent? Br.

J. Pharmacol. 64:566–574. https://doi.org/10.1111/j.1365-2125.2007.03021.x.

Thanacoody, R.B.H.K., 2011. Thioridazine: the good and the bad. Recent Pat. Antiinfect.

Drug Discov. 6, 92 –98.

Trautwein, C., Kümmerer, K., 2012a. Degradation of the tricyclic antipsychotic drug

chlorpromazine under environmental conditions, identiﬁcation of its main

aquatic biotic and abiotic transformation products by LC-MSn and their effects

on environmental bacteria. J. Chromatogr. B 889–890:24–38. https://doi.org/

10.1016/j.jchromb.2012.01.022.

Mahmoud, W.M.M., Toolaram, A.P., Menz, J., Leder, C., Schneider, M., Kümmerer, K., 2014.

Identiﬁcation of phototransformation products of thalidomide and mixture toxicity as-

sessment: an experimental and quantitative structural activity relationships (QSAR)

approach. Water Res. 49:11–22. https://doi.org/10.1016/j.watres.2013.11.014.

Manju, T., Manoj, N., Braun, A.M., Oliveros, E., 2012. Self-sensitized photooxidation of N-

methyl phenothiazine: acidity control of the competition between electron and ener-

gy transfer mechanisms. Photochem. Photobiol. Sci. 1744. https://doi.org/10.1039/

c2pp25244a.

Menz, J., Schneider, M., Kümmerer, K., 2013. Toxicity testing with luminescent bacte-

ria — characterization of an automated method for the combined assessment of

acute and chronic effects. Chemosphere 93:990–996. https://doi.org/10.1016/

j.chemosphere.2013.05.067.

Trautwein, C., Kümmerer, K., 2012b. Ready biodegradability of triﬂuoromethylated phe-

nothiazine drugs, structural elucidation of their aquatic transformation products,

and identiﬁcation of environmental risks studied by LC-MSⁿand QSAR. Environ. Sci.

Pollut. Res. 19:3162–3177. https://doi.org/10.1007/s11356-012-1002-1.

USEPA, 2012. U.S. EPA. Environmental protection agency EHAC. PBT proﬁler: a compo-

nent of OPPT's P2 Frame work

V 2.000; 2012. [WWW Document]. http://

www.pbtpro ﬁ ler.net/, Accessed date: 31 March 2015.

Vandermeersch, G., Lourenço, H.M., Alvarez-Muñoz, D., Cunha, S., Diogène, J., Cano-

Sancho, G., Sloth, J.J., Kwadijk, C., Barcelo, D., Allegaert, W., Bekaert, K., Fernandes,

J.O., Marques, A., Robbens, J., 2015. Environmental contaminants of emerging concern

in seafood — European database on contaminant levels. Environ. Res. 1–17. https://

doi.org/10.1016/j.envres.2015.06.011.

Please cite this article as: Wilde, M.L., et al., Combination of experimental and in silico methods for the assessment of the phototransformation

products of the antipsychotic drug/metabolite Mesoridazine, Sci Total Environ (2017), https://doi.org/10.1016/j.scitotenv.2017.08.040

M.L. Wilde et al. / Science of the Total Environment xxx (2017) xxx–xxx

Wammer, K.H., Korte, A.R., Lundeen, R.A., Sundberg, J.E., McNeill, K., Arnold, W.A., 2013. Di-

rect photochemistry of three ﬂuoroquinolone antibacterials: norﬂoxacin, oﬂoxacin, and

enroﬂoxacin. Water Res. 47:439–448. https://doi.org/10.1016/j.watres.2012.10.025.

Wang, X.H., Lin, A.Y.C., 2014. Is the phototransformation of pharmaceuticals a natural pu-

riﬁcation process that decreases ecological and human health risks? Environ. Pollut.

186:203–215. https://doi.org/10.1016/j.envpol.2013.12.007.

Werner, J.J., Arnold, W.A., Mcneill, K., 2006. Water hardness as a photochemical parame-

ter: tetracycline photolysis as a function of calcium concentration, magnesium con-

centration, and pH. Environ. Sci. Technol. 40:7236–7241. https://doi.org/10.1021/

es060337m.

Wilde, M.L., Menz, J., Trautwein, C., Leder, C., Kümmerer, K., 2016. Environmental

fate and effect assessment of thioridazine and its transformation products

formed by photodegradation. Environ. Pollut. 213:658–670. https://doi.org/

10.1016/j.envpol.2016.03.018.

Wilde, M.L., Schneider, M., Kümmerer, K., 2017. Fenton process on single and mixture

components of phenothiazine pharmaceuticals: assessment of intermediaries, fate,

and preliminary ecotoxicity. Sci. Total Environ. 583:36–52. https://doi.org/10.1016/

j.scitotenv.2016.12.184.

neuroleptic thioridazine. Drug Metab. Dispos. 34:471. https://doi.org/10.1124/

dmd.105.006445.tion.

Wu, X., Conkle, J.L., Ernst, F., Gan, J., 2014. Treated wastewater irrigation: uptake of phar-

maceutical and personal care products by common vegetables under field conditions.

Environ. Sci. Technol. 48, 11286 –11293.

Zepp, R.G., 1978. Quantum yields for reaction of pollutants in dilute aqueous solution. En-

viron. Sci. Technol. 12:1976–1978. https://doi.org/10.1021/es60139a010.

Zepp, R.G., Cline, D.M., 1977. Rates of direct photolysis in aquatic environment. Environ.

Sci. Technol. 11:359–366. https://doi.org/10.1021/es60127a013.

Zhu, B., Zonja, B., Gonzalez, O., Sans, C., Pérez, S., Barceló, D., Esplugas, S., Xu, K., Qiang, Z.,

2015. Degradation kinetics and pathways of three calcium channel blockers under UV

irradiation. Water Res. 86:9–16. https://doi.org/10.1016/j.watres.2015.05.028.

Zonja, B., Pérez, S., Barceló, D., 2016. Human metabolite lamotrigine-N2-glucuronide is

the principal source of lamotrigine-derived compounds in wastewater treatment

plants and surface water. Environ. Sci. Technol. 50:154–164. https://doi.org/

10.1021/acs.est.5b03691.

Wójcikowski, J., Maurel, P., Daniel, W.A., 2006. Characterization of human cytochrome

p450 enzymes involved in the metabolism of the piperidine-type phenothiazine

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