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New Safe Method for Preparation of Sarin-Exposed Human Erythrocytes Acetylcholinesterase Using Non-Toxic and Stable Sarin Analogue Isopropyl p-Nitrophenyl Methylphosphonate and its Application to Evaluation of Nerve Agent Antidotes

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Abstract

Introduction

A non-toxic and stable sarin analogue, isopropyl p-nitrophenyl methylphosphonate (INMP), was synthesized for safe preparation of sarin-exposed acetylcholinesterase (AChE).

Results and Discussion

This agent was stable for years, able to be handled in an ordinary laboratory without special care, and its 50% inhibitory concentration (IC50) on 0.04 U/ml human erythrocytes AChE was 15 nM. This reagent was thought to be especially useful since it enables experiments that require sarin-inhibited AChE, such as the development of antidotes for sarin, in a usual laboratory. To demonstrate the usefulness of this method, 40 known and novel pyridinealdoxime methiodide (PAM)-type oxime antidotes were synthesized, and their reactivation activities to INMP-exposed AChE and structure–activities correlation were studied.

Conclusion

Among the antidotes tested in this experiment except for 2-PAM, the compound found to have the highest reactivation activity, was the novel hydrophobic 2-PAM-type compound, 2-[(hydroxyimino)methyl]-1-[4-(tert-butyl)benzyl] pyridinium bromide.

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Abbreviations

AChE:

acetylcholinesterase

BBB:

blood–brain barrier

BIMP:

bis(isopropyl methylphosphonate)

CH2Cl2 :

dichloromethane

DCC:

dicyclohexylcarbodiimide

DCU:

dicyclohexylurea

DMAP:

dimethylaminopyridine

IC50 :

50% inhibitory concentration

IMPA:

methylphosphonic acid monoisopropyl ester

INMP:

isopropyl p-nitrophenyl methylphosphonate

LC–ESI-MS:

liquid chromatography–electrospray mass spectrometry

NMR:

nuclear magnetic resonance

PAM:

pyridinealdoxime methiodide

pK a :

dissociation constant

RT:

room temperature

THF:

tetrahydrofuran

TLC:

thin-layer chromatography

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Acknowledgment

This study was supported in part by a Grant-in-Aid for Scientific Research (B) from the Ministry of Education, Culture, Sports, Science and Technology of Japan.

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Authors and Affiliations

  1. Toxicology Section, Department of Forensic Chemistry, National Research Institute of Police Science, 6-3-1, Kashiwa-no-ha, Kashiwa, Chiba, 277-0882, Japan

    Hikoto Ohta

  2. Department of Forensic Biology, National Research Institute of Police Science, 6-3-1, Kashiwa-no-ha, Kashiwa, Chiba, 277-0882, Japan

    Hiroshi Ikegaya & Koichi Sakurada

  3. National Research Institute of Police Science, 6-3-1, Kashiwa-no-ha, Kashiwa, Chiba, 277-0882, Japan

    Takehiko Takatori

  4. Chemical Warfare Section, Department of Forensic Chemistry, National Research Institute of Police Science, 6-3-1, Kashiwa-no-ha, Kashiwa, Chiba, 277-0882, Japan

    Takeshi Ohmori

  5. Physical Evidence Section, Department of Forensic Chemistry, National Research Institute of Police Science, 6-3-1, Kashiwa-no-ha, Kashiwa, Chiba, 277-0882, Japan

    Shinichi Suzuki

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  1. Hikoto Ohta
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  2. Takeshi Ohmori
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  3. Shinichi Suzuki
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  4. Hiroshi Ikegaya
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  5. Koichi Sakurada
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  6. Takehiko Takatori
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Correspondence to Hikoto Ohta.

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Ohta, H., Ohmori, T., Suzuki, S. et al. New Safe Method for Preparation of Sarin-Exposed Human Erythrocytes Acetylcholinesterase Using Non-Toxic and Stable Sarin Analogue Isopropyl p-Nitrophenyl Methylphosphonate and its Application to Evaluation of Nerve Agent Antidotes. Pharm Res 23, 2827–2833 (2006). https://doi.org/10.1007/s11095-006-9123-1

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  • DOI: https://doi.org/10.1007/s11095-006-9123-1

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