Inhibition of Acetylcholinesterase Activity by Volatile α, β-Unsaturated Ketones

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  • This article was downloaded by: [University of California Davis]On: 09 October 2014, At: 18:50Publisher: Taylor & FrancisInforma Ltd Registered in England and Wales Registered Number: 1072954 Registeredoffice: Mortimer House, 37-41 Mortimer Street, London W1T 3JH, UK

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    Inhibition of AcetylcholinesteraseActivity by Volatile , -UnsaturatedKetonesMitsuo Miyazawa a , Aya Kakiuchi a , Hitomi Watanabe a & HiromuKameoka aa Department of Applied Chemistry, Faculty of Science andEngineering , Kinki University , Kowakae, Higashiosaka-shi, Osaka,577-8502, JapanPublished online: 04 Oct 2006.

    To cite this article: Mitsuo Miyazawa , Aya Kakiuchi , Hitomi Watanabe & Hiromu Kameoka (1998)Inhibition of Acetylcholinesterase Activity by Volatile , -Unsaturated Ketones, Natural ProductLetters, 12:2, 131-134, DOI: 10.1080/10575639808048281

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  • Natural Product Leiters Volume 12(2), pp. 131-134 Reprints available directly from the Publisher Photocopying permitted by license only

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    Inhibition of Acetylcholinesterase Activity by Volatile a, &Unsaturated Ketones

    MlTSUO MIYAZAWA,* AYA KAKIUCHI, HITOMI WNANABE and HIROMU KAMEOKA

    Department of Applied Chemistry, Faculty of Science and Engineering, JSinki University, Kowakae, Higashiosaka-shi, Osaka 577-8502, Japan

    (Received 2nd March 1998)

    Abstract: Inhibition of acetylcholinesterase (AChE) activity by 5 kinds of Inhibition of AChE was measured volatile a, p-unsaturated ketones was studied.

    by the colorimetric method. d o n o n e showed the strongest inhibition.

    Key word: Acetylcitohesteulre; vo lde a, /I- un.satum&d ketones; inhibdion of enzyme activity; a-wmne

    INTRODUCTION The antimicrobial activity of essential oils is well recognized. It is reported

    thatcarvacrol, c i t d and geraniol of essential oils have antibacterial activity against SalmonelIa fyphurium', and gowth inhibition of Leukemia Cell Line CEM-C1 by farnesol. * As part of continuing program to search for the bioactive natural compounds, we

    investigated the inhibition of acetylcholinesterase (AChE) activity by volatile a, p- unsaturated ketones. The volatile unsaturated ketones are existing in essential oils of plants. There are only three reports about inhibition of AChE activity from the housefly and electric eel by m~noterpenoids.~" In our previous pap#, the inhibition of A C E of mammals by monoterpenoids having a p-menthane skeleton was reported. In the present paper, we report the inhibition of AChE from bovine erythrocytes by 5 kinds of volatile a, f3-unsaturated ketones and their structure- activity relationships were also discussed.

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  • 132 M. MIYAZAWA et al.

    MATERIALS AND METHODS Materials: AChE from bovine erythrocytes was purchased from Seikagaku

    Kogyo Co., Ltd. (Tokyo). 5,S-Dithiobis (2-nitrobenzoic acid) (DTNB) was purchased from Tokyo Kasei Kogyo Co.. Ud. (TCI) (Tokyo). Acetylthiocholine iodide (ATC) was purchased from Kanto Chemical Co., Inc. (Tokyo). Volatile a, 6- unsaturated ketones were purchased from Yarnamoto Perfume Co., Ltd. (Osaka). Preparatory Solutions: AChE(0.04 U/ml) and ATC (75 mL) were dissolved

    in 0.1 M phosphate buffer (pH 8.0, respectively). DTNB (0.01 M) was made up in 10 mL of 0.1 M phosphate buffer (pH 7.0) containing 15 mg of NaHCQ. Volatile a, b- unsaturated ketones in all assays were maintained at 5 % (v/v), including controls.

    Inhibition of AChE was assessed by the colorimetric method of Ellman7 weremixed in a test tube, and the tube was set on the incubator (25 "C). To the tube were added DTNB (100 pL) and buffer (2.4 mL), and the reaction was started by adding ATC (40 pL). The mixture was then incubated at 25 C for 5 min as preincubation. The reaction was started by adding ATC (40 pL), and the mixture was incubated at 25 "C for 20 min. The absorbance at 412 nm was measured specfrophotometrically, and all test and control (without volatile a, B- unsaturated ketone) assays were corrected by blanks for nonenzymic hydrolysis. Each assay was run in triplicate, at a minimum.

    Inhibition of AChE Activity: Inhibitor solution (50 pL) and AChE (0.5 ml)

    RESULTS AND DISCUSSION The dose dependent inhibition of AChE activity by 5 kinds of volatile a , f3- unsaturated ketones (Figure 1) treatment, is shown in Figure 2. The AChE activity values for the inhibitors were calculated as compared to the control (without volatile a, p-unsaturated ketones) AChE activity, assumed to be 100%. Table 1 shows 50% enzyme inhibition (ICso) and percentage of inhibition of AChE activity (%) at 80 pg/ml calculated from the inhibition dose-response curve. As shown in Figure 2, cis-jasmone (2), p-methyl naphthyl ketone (3), 6-ionone (4) and a-ionone ( 5 ) showed the inhibition ofAChE activity from 53% to 71% at 80 pg/ml, and IC,, values were 78.3, 55.0, 53.3, and 36.7 @ml, respectively.

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  • 133 INHIBITN. ACHE-ACTIVITY BY VOLATILE a, P-UNSATD. KETONES

    Acetophenone (1) inhibited 17.8 % , and no IC,, value was obtained at < 80 Wgjml. With regard to isomer 4 and 5, 5 showed stronger inhibition than 4. On the other hand, the aliphatic compounds(4 and 5) showed stronger inhibition than the aromatic compounds(1 and 3). Also, 3 which has a naphthalene ring showed stronger inhibition than 1 has a benzene ring.

    p-methyl naphthyl ketone (3) 0 50 100

    Inhibition Concentration (&mI)

    Figure 2. Bfect of volatile a. b- unsaturated ketones on A C E activity. The percentage of enzyme activity values for the inhibitor was calculated as compared to

    p-ionone (4) a-ionone (5) the control activity, assumed to be 100%. (0) pulegone, (0) acetophenone (I). (0) cis-jasmone (2). (A) &methyl naphthyl ketone (3). (m) B-ionone (4).

    PP F i V 1. structures of volatile a, p- unsaturated

    ketones. (*) a-ionone (5).

    Table 1. Inhibition of AChE by Volatile a, f3- Unsaturated Ketones.

    Chemicals Icso (p.g/ml)a Inhibition activity (%)b

    1 - 2 78.3 3 55.0 4 53.3 5 36.7

    17.8 53.1 65.8 60.3 70.3

    Conocntration of chemicals required for 5PA enzyme inhibition as calculated from the dose-response curve.

    The percent AChE inhibition values (80 &nl) were calculated as compared to control (without volatile a, fl- unsaturated ketones) enzyme activity (assumed to be

    0% inhibition).

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  • 134 M. MIYAZAWA et al.

    ACKNOWLEDGMENT

    We are grateful to the Environmental Science Institute of Kinki University for financial support.

    LITERATURE CITED

    * To whom correspondence should be addressed. (fax +81-6-727-4301; E-mail:

    1. Kim, M. J., Marshall, R. M., Cornell. A. J., Preston 111, F. J . , and Wel, C. I . ,

    2. Melnykovych, G.,. Haug, J. S. and Goldner, C. M. (1992) Bwchem. Biophys.

    3. Gracza, L. (1961) Biochem. Phamrazie., 7, 88. 4. Grundy, D. L., and Stll, C. C. (1985) Pestic. Biochem. Physiof., 23, 383. 5. Ryan, M. F. and Byme, 0. (1988) J. Chem. Ecol., 14, 1965. 6. Miyazawa, M, Watanabe, H., and KameokaH. (1997) J. Agric. Food Chem.,

    7. Ellman, G. L, Courtney, K. D., Valentino. A. Jr. and Feathertone, R. M.

    miyazawa @ apch. kindai. ac. jp)

    (1995) J. Food Sci., 6 0, 1364.

    Res. Commun., 186, 543.

    45,677.

    (1961) Biochem. P h a m i e . , 7 , 88.

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