3.13.1
© Springer-Verlag Berlin Heidelberg 2005
II.3.1 Phenothiazines
by Akira Ishii and Yoshinao Katsumata
Introduction
Phenothiazine drugs, including chlorpromazine and levomepromazine, have been being
widely used as neuroleptics ( major tranquilizers), antiparkinsonian drugs and antihistaminics
for a long time [1].
> Table 1.1 shows chemical structures of representative phenothiazines.
 ese drugs show blocking action on D
2
receptors of dopaminergic neurons; there is close
relationship between the receptor blocking and tranquilizing actions.  e dopamine D
2
recep-
tor-blocking actions provoke extrapyramidal symptoms, such as muscular sti ness, tremor
and ptyalism. Orthostatic hypotension, arrythmia and icterus are occasionally found a er ad-
ministration of phenothiazines as side e ects.
 e number of phenothiazine poisoning cases is relatively small, as compared with the
extensive use of this group of drugs [2]. However, until now, many phenothiazine poisoning
cases, including fatal ones [3, 4], were reported.
Phenothiazines were analyzed by various methods, such as GC, GC/MS, HPLC and LC/
MS.  e methods by GC and GC/MS are relatively simple, but most of the methods reported
used packed columns or wide-bore capillary columns [5, 6], which did not give high sensi-
tivity.
Hattori et al. [7] reported a highly sensitive method for detecting phenothiazine in body
 uids using GC- surface ionization detection ( SID)
a
. Although the SID detector has an advan-
tage in that each compound having a tertiary amino group can be analyzed with high sensitiv-
ity and speci city, it is not a detector commonly available. In this chapter, the methods

GC column: an Rtx-1 fused silica capillary column (30 m × 0.32 mm i.d.,  lm thickness
0.25 µm, Restek, Bellefonte, PA, USA)
c
GC conditions; instrument: a Shimadzu GC-17A gas chromatograph connected to MS
(Shimadzu Corp., Kyoto, Japan)
d
; column (oven) temperature: 150 °C (1 min) →15 °C/min
e
→
290 °C (10 min); injection temperature: 270 °C; interface temperature; 270 °C; carrier gas: He;
its  ow rate: about 1.5 mL/min; injection: splitless mode for 1 min followed by the split mode.
MS conditions; instrument: a Shimadzu QP-5050A quadrupole mass spectrometer
f
; mea-
surement: scan mode; ionization: positive ion EI; ionization current: 60 µA; electron energy:
70 eV; scan range: m/z 50–400; scan speed: 1,000 amu/s.
Procedure
i. A 10-mL volume of methanol and 10 mL distilled water are passed through a Sep-Pak C
18

cartridge; this procedure is repeated twice (in total 3 times) to activate the cartridge
g
.
ii. To 1 mL of whole blood or urine, are added 8 mL distilled water
h
, internal standard (IS)
solution and 1 mL of 1 M sodium bicarbonate solution. For IS, a non-target phenothiazine
drug (200 ng)
i
is selected for use.

.
 e GC/MS analysis for phenothiazines shows some disadvantages as follows. ① Pheno-
thiazines with long side chains are not suitable for GC/MS analysis; especially, those with long
piperazinyl or hydroxyl side chains cannot be detected by either GC or GC/MS. ② Prometha-
zine, isothipendyl and ethopropazine do not give molecular ions, but give big fragment peaks
at m/z 58 or 72 only, which are not useful for identi cation of the compounds. For analysis of
such compounds, the following LC/MS becomes very useful.
LC/MS analysis [8]
See [8].
Reagents and their preparation
i. Reagents
Prochlorperazine, tri uoperazine, perphenazine,  uphenazine, propericiazine and thiorida-
zine can be purchased from Sigma (St. Louis, MO, USA). Pure powder of perazine and clospi-
razine was donated by Mitsubishi Welpharma, Osaka, Japan; that of  upentixol by Takeda
Chem. Ind. Ltd., Osaka, Japan; that of thioproperazine by Shionogi & Co., Ltd., Osaka, Japan;
that of thiethylperazine by Sandoz, Basel, Switzerland.
ii. Preparation
 e above compounds are all in the salt forms; 1 mg free base/mL methanolic solution is pre-
pared for each compound.
LC/MS conditions
LC column: a Capcell Pak C
18
UG-80 capillary column (S-5 µm, 250 × 1.0 mm i.d., Shiseido,
Tokyo, Japan).
LC conditions; instrument
l
: an HP-1100 Series LC chromatograph (Agilent Technologies,
Palo Alto, CA, USA); mobile phase A: distilled water containing 0.1 % formic acid and 10 mM
259
ammonium acetate; mobile phase B: 100 % acetonitrile. A gradient elution with solutions A

whole blood and urine cannot be analyzed by LC/MS. To overcome this di culty, detection
using LC/tandem MS seems useful. For the details of the tandem MS for phenothiazines, the
readers can refer to the reference [8]. According to the reference, every phenothiazine with a
long side chain can be speci cally detected with a high S/N ratio. A report dealing with the
combination of LC/MS/MS with solid-phase microextraction ( SPME) for phenothiazines was
also published [9].
LC/MS analysis