A highly selective visual detection of tabun mimic diethyl cyanophosphate (DCNP): Effective discrimination of DCNP from other nerve agent mimics

Article abstract of DOI:10.1039/c4ra02742f

A new tabun specific visual detection protocol is reported. The chemodosimeter fluorescein-hydroxamate aldehyde undergoes tandem nucleophilic substitution followed by cyanohydrin reaction with DCNP and shows a specific spectroscopic behaviour. The probe molecule clearly distinguishes tabun mimic DCNP over other nerve agent simulants as well as strong electrophiles.

Full text of DOI:10.1039/c4ra02742f

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A highly selective visual detection of tabun mimic
diethyl cyanophosphate (DCNP): effective
discrimination of DCNP from other nerve agent
mimics†
Cite this: RSC Adv., 2014, 4, 24645
Received 28th March 2014
Accepted 22nd May 2014
D. Raghavender Goud, Deepak Pardasani, Vijay Tak and Devendra Kumar Dubey*
DOI: 10.1039/c4ra02742f
www.rsc.org/advances
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A new tabun specific visual detection protocol is reported. The che- containing sensors, nucleophilic substitution reactions and
15
modosimeter fluorescein-hydroxamate aldehyde undergoes tandem complex formation based probes. The main drawback of these
nucleophilic substitution followed by cyanohydrin reaction with DCNP methods is lack of specicity towards the target analyte. Most of
and shows a specific spectroscopic behaviour. The probe molecule these methods mainly target the electrophilicity of nerve agents,
clearly distinguishes tabun mimic DCNP over other nerve agent sim- therefore the other reactive electrophiles also give the similar
ulants as well as strong electrophiles.
response as nerve agents. Although the disaster response
protocol is similar for all nerve agents, but the choice of oxime
for AChE enzyme reactivation is not same. Experimental
evidences shows that PAM is preferred for sarin and VX, HI-6 for
soman and obidoxime for tabun. These reasons emphasise the
need for the development of specic detection methods for each
nerve agent.
Herein, we report a tabun specic chemodosimeter molecule
that is able to differentiate DCNP (tabun mimic) from other
nerve agent mimics as well as strong electrophiles. The main
structural difference among the tabun and other nerve agents or
their mimics is the electrophilic phosphorous centre is bonded
to cyanide anion in tabun. Whereas in case of other nerve
agents the anions other than cyanide like uoride, thiolate and
chloride were bonded to electrophilic phosphorous centre of
the nerve agents (Fig. 1). Therefore based on these observations
we expected that specic colorimetric detection of the tabun is
possible by targeting both the electrophilic character and
cyanide anion present in tabun. In this study we have chosen
DCNP as a tabun mimic as it behaves similar to tabun due to the
Nerve agents are the most toxic of the known chemical warfare
agents. These agents are hazardous and can cause death within
minutes aer exposure. The ease of production, extreme toxicity
and the current rise in international concern over the use of
chemical warfare agents in terrorism, accentuates the need to
develop simple and reliable methods to detect these lethal
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1
chemicals. Nerve agents inhibit the enzyme acetylcholines-
terase (AChE), which degrades the neurotransmitter acetylcho-
line. This leads to accumulation of acetylcholine at nerve
synapses resulting neuromuscular paralysis and eventually
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death. Various methods for the detection of nerve agents have
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been developed including enzymatic assays, optical-ber
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arrays, surface acoustic wave (SAW) devices, electrochemistry,
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biosensors and ion mobility spectroscopy. However, they show
at least one of the following limitations; operational complexity,
non-portability, high cost, difficulties in real-time monitoring
and false positive readings.
In recent years development of chromogenic and uores-
cence detection methods have gained considerable interest as
an alternative to above mentioned complex methods. The main
advantages of chromogenic detection method are portability,
easy to operate, cheap and capable of real time detection. The
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recently developed methods are PET based probes, cyclisation
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reactions in push–pull chromophores, displacement like
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assays,
molecular impregnated polymers,
oximate
Vertox Laboratory, Defence Research and Development Establishment, Jhansi Road,
Gwalior, M.P. 474002, India. E-mail: dkdubey@rediffmail.com; Fax: +91-0751-
2
341148
†
Electronic supplementary information (ESI) available: Experimental details and
additional characterization data. See DOI: 10.1039/c4ra02742f
Fig. 1 Structures of nerve agents and their mimics.
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presence of electrophilic phosphorous centre is attached to
cyanide anion but less toxic than tabun.
The tabun specic chemodosimeter molecule uorescein-
hydroxamate aldehyde (I) was designed in such a way that it
consist of two reactive sites (Scheme 1). One is oximate group
that acts as a nucleophile and is reactive towards the electro-
philic phosphorous centre of the nerve agents. Another reactive
site is aldehyde group that acts as an electrophile and selectively
reacts with the cyanide anion released from DCNP via cyano-
hydrin formation. This cascade reaction may result in the
specic optical behaviour towards DCNP.
The probe molecule was synthesized by a three step proce-
dure. In the rst step uorescein methyl ester was synthesized presence of DCP and DCNP (10 equivalents).
by Fischer esterication of uorescein. In the next step uo-
Fig. 2 UV/Vis spectra of chemodosimeter (I) (C ¼ 1 mM) solution in the
rescein hydroxamate was synthesized by nucleophilic substitu-
tion with hydroxylamine hydrochloride. Finally the target nucleophilic substitution to give an intermediate (IIA). In the
molecule was synthesized by introducing the aldehyde group by next step the intermediate (IIA) undergoes Lossen rearrange-
Riemer–Tiemann reaction (Scheme 1).
ment to afford conjugated product (IIIA), which is responsible
To test selectivity of the probe molecule, the reactions of for the bathochromic shi to 530 nm (Fig. 2). Similarly by the
probe molecule with DCNP (tabun mimic) and DCP (nerve agent addition of tabun mimic DCNP (10 equivalents) to the test
mimic) were screened in various solvents. Among these DMF solution, colour change to blue-green was observed. When
and DMSO were optimized for DCP and DCNP respectively. The DCNP reacts with the probe molecule, the hydroxamate group
optimized test solution consists of the chemodosimeter (1 mg of the probe molecule reacts at electrophilic phosphorous
ꢀ1
ml ) and 3% (v/v) triethylamine as a base. With DCP the probe centre of DCNP and subsequently the released cyanide anion
molecule gives intense red colour in DMF, where as in DMSO it undergoes electrophilic addition reaction (cyanohydrin forma-
gives weak orange-red colour. This may be due to the negative tion) with aldehyde group to give intermediate (IIB). In the next
17
solvatochromic effect. Therefore better sensitivity can be step the intermediate undergoes Lossen rearrangement to
achieved for DCP in DMF when compared to DMSO. Due to this result in the formation of another highly conjugated product
reason DMF was preferred for DCP analysis. In case of DCNP (IIIB) and there was a further bathochromic shi to 608 nm
5
mg of KI was used as a catalyst. As DCNP is less reactive than (Fig. 2) was observed which is responsible for the blue-green
DCP, iodide by virtue of its polarisable nucleophilicity and colour. In product (IIIA) the delocalisation of conjugated elec-
nucleofugicity facilitates the phosphorylation of (I) by efficiently trons was retarded due to the presence of highly electron
knocking out the cyanide present in DCNP. KI doesn't interfere withdrawing aldehyde group (Scheme 2).
with the detection of DCP. Initially the test solutions were yellow
Where as in case of product (IIIB) due to the formation of
in colour and exhibit a unique band at 410 nm (Fig. 2). When a cyanohydrin the electron withdrawing effect of aldehyde group
nerve agent simulant DCP (10 equivalents) was added to the test is lost, this increases the degree of delocalisation in the product
solution a remarkable colour modulation from yellow to red was (IIIB) which may be responsible for further bathochromic shi.
observed. When DCP reacts with the chemodosimeter mole- The reaction of uorescein-hydroxamate aldehyde with DCNP
cule, the hydroxamate group of the probe molecule reacts at and DCP was monitored by subjecting the reaction solution to
electrophilic phosphorous centre of the nerve agent mimic via ESI-MS analysis. The peaks corresponding to the molecular
weight of the intermediates and the products were identied,
this conrms the formation of these products in assay systems
(See ESI, Fig. S6 and S7†).
In the next step, to test the interferences the chemo-
dosimeter molecule was reacted with various strong electro-
philes like thionyl chloride, phosphorous oxychloride, 4-
toluenesulfonyl chloride and acetyl chloride. In all the cases
instantaneous colour change to red was observed similar to
DCP as shown in the Fig. 3. In addition to this, interferences of
anions like cyanide, iodide, bromide and uoride were also
checked. No change in spectroscopic behaviour was noted in
the presence of any of these anions (Fig. 3). Only with DCNP (10
equivalent) specic colour change to blue-green was observed
(Fig. 3), which indicates that the probe molecule clearly
distinguishes the tabun mimic DCNP over other electrophiles
and nerve agent mimics. The chemodosimeter (I) was also
checked for uorescence response against DCP and DCNP. The
Scheme 1 Synthesis of fluorescein-hydroxamate aldehyde (I).
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formation of intricate reaction mixture. The chromogenic
response was found to be reproducible for both the analytes.
In summary, a visual and highly selective detection method
for tabun mimic DCNP was developed by targeting the electro-
philicity as well as the presence of cyanide anion in DCNP. The
probe molecule uorescein-hydroxamate aldehyde undergoes
tandem phosphorylation followed by cyanohydrin reaction with
DCNP to afford a specic product, which shows the specic
spectroscopic behaviour. This specic product formation takes
place only in the presence of DCNP, therefore no false positives
were noted for all possible closely related interferences. We
have successfully demonstrated the possibility of detecting
tabun mimic DCNP using a tandem reaction based probe
molecule. The similar approach can be used to develop the
probe molecules for the selective detection of other nerve
agents.
ꢀ
1
Fig. 3 Chromogenic response of chemodosimeter (I) (1 mg ml
)
,
solutions: from left to right I, I + DCP, I + DCNP, I + SOCl
2
, I + POCl
3
ꢀ
ꢀ
ꢀ
ꢀ
I + C
7
H
8
SO
2
Cl, I + CH
3
COCl, I + CN , I + I , I + Br , I + F
(
10 equivalents each).
probe molecule was found to be uorescent for DCP but non-

uorescent for DCNP.
To test the sensitivity of the chemodosimeter towards tabun
mimic DCNP and nerve agent mimic DCP, the analytes with
various concentrations were added to their respective reagent
solutions. In case of DCP the colour change to red was instan- Notes and references
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Scheme 2 Chromogenic response mechanism for chemodosimeter
(I) in the presence of nerve agent simulants DCP and DCNP.
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