Syntheses of isocitric acid derivatives and biological evaluation

Article abstract of DOI:10.1016/S0040-4039(00)00316-6

Isocitric acid methyl esters and their derivatives have been synthesized and their biological activities, superoxide release inhibition and tumor necrosis factor-alpha (TNFα)'s release inhibition, have been studied. Linoleic and linolenic acid derivatives showed strong activity against TNFα's release inhibition comparable to sarcophytol A and cryptoporic acid E. (C) 2000 Elsevier Science Ltd.

Full text of DOI:10.1016/S0040-4039(00)00316-6

TETRAHEDRON
LETTERS
Pergamon
Tetrahedron Letters 41 (2000) 3095–3098
Syntheses of isocitric acid derivatives and biological evaluation
Motoo Tori,a,∗ Yoko Sono,a Yasuyo Nakashiba,a Noriko Hamada,a Masakazu Sono,a
Yoshinori Asakawa,a,∗ Masami Suganuma,b Sachiko Okabe b and Hirota Fujiki b
aFaculty of Pharmaceutical Sciences, Tokushima Bunri University, Yamashiro cho, Tokushima 770-8514, Japan
bSaitama Cancer Center Research Institute, Ina, Kitaadachi-gun, Saitama 362-0806, Japan
Received 17 January 2000; accepted 17 February 2000
Abstract
Isocitric acid methyl esters and their derivatives have been synthesized and their biological activities, superoxide
release inhibition and tumor necrosis factor-alpha (TNFα)’s release inhibition, have been studied. Linoleic and
linolenic acid derivatives showed strong activity against TNFα’s release inhibition comparable to sarcophytol A
and cryptoporic acid E. © 2000 Elsevier Science Ltd. All rights reserved.
Keywords: biologically active compounds; carboxylic acids and derivatives; esters; terpenes and terpenoids.
We have previously reported the isolation of cryptoporic acids A–F from the fungus Cryptoporus
volvatus and established their structures based on high resolution 2D NMR techniques, X-ray analysis,
and chemical degradations.1–5 It is very interesting to note that cryptoporic acids exhibit strong supero-
xide release inhibition and anti-tumor promotion activities.5 The structures are unusual; namely, they are
drimane-type sesquiterpenes linked to isocitric acid moiety with an ether bond.6–10 We were interested
in their biological activities and planned to prepare synthetic substitutes to the natural products.11 We
first intended to prepare isocitric acid with the same stereochemistry as natural products. Then fatty acids
were attached to the isocitric acid and biological activities of these esters were evaluated. We now report
our preliminary results.
Since Seebach et al. reported the alkylation of malic acid and synthesis of isocitric acid,12 we have
modified their methodology to prepare monocarboxylic acid derivatives13 (Scheme 1). The allylated
compound 2 of (R)-(+)- or racemic malic acid was protected with the MOM group and subjected to
ozonolysis. The Jones’ oxidation of aldehyde 4 afforded acid 5, which was converted to trichloroethyl
ester 6 using DCC. The MOM group was deprotected by TMSBr and fatty acids were used for the
preparation of esters, whose trichloroethyl group was removed under Zn–AcOH–MeOH conditions to
afford 8, 9 and 10.14 Trimethyl isocitrate (11)14 was synthesized by methylation (CH2N2) of acid 5 and
deprotection of the MOM group in 65% yield from 5.
∗
Corresponding authors. Tel: +81-88-622-9611; fax: +81-88-655-3051; e-mail: tori@ph.bunri-u.ac.jp (M. Tori), asakawa@
ph.bunri-u.ac.jp (Y. Asakawa)
0040-4039/00/$ - see front matter © 2000 Elsevier Science Ltd. All rights reserved.
PII: S0040-4039(00)00316-6
tetl 16596

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Scheme 1. Reagents and conditions: (a) LDA, BrCH2CH_CH2; (b) MOM–Cl, EtNiPr2; (c) O3, CH2Cl2–MeOH, then SMe2;
(d) Jones; (e) DCC, HOCH2CCl3, DMAP; (f) BrSiMe3, CH2Cl2; (g) RCOOH, DCC, DMAP, CH2Cl2; (h) Zn, AcOH, MeOH
Malic acid derivatives were similarly prepared (Scheme 2). The MOM-protected methyl glycolate was
allylated, and similarly converted to methyl trichloroethyl ester of the malic acid 15. The esterification
with fatty acids using DCC and deprotection afforded 16 and 17.14
Scheme 2. Reagents and conditions: (a) LDA, BrCH2CH_CH2; (b) O3, CH2Cl2–MeOH, then SMe2; (c) Jones; (d) DCC,
HOCH2CCl3, DMAP; (e) BrSiMe3, CH2Cl2; (f) RCOOH, DCC, DMAP, CH2Cl2; (g) Zn, AcOH, MeOH
Because the natural products exhibit strong superoxide release inhibition activities,4,5 these compounds
were subjected to the same tests.4,5 The results are listed in Table 1.15 Among those tested, compound
16 was the most effective (IC50 0.39 µM). The malic acid derivatives were 10 times more effective than
isocitric acid derivatives. The linoleic acid derivatives were more than twice as strong as linolenic acid
derivatives.
Compounds 8 and 9 were further subjected to inhibition tests of TNFα’s release with okadaic acid
by anti-tumor promoters on BALB/3T3 cells.5,16 As shown in Fig. 1, both compounds are as strong as
sarcophytol A (18) and cryptoporic acid E (19). Therefore, it is very interesting to note that the relative
positions of the hydrophilic and lipophilic parts are very important, and that the lipophilic part can be
substituted by fatty acids to show anti-tumor promoter activity.
Thus, we have prepared fatty acid derivatives with isocitric and malic acid moiety and evaluated their
biological activities, and two of them (8 and 9) were as effective as sarcophytol A (18) and cryptoporic
acid E (19) in inhibition of TNFα’s release.

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Table 1
Superoxide release inhibition activity against guinea pig macrophage
Fig. 1. Inhibition of TNFα’s release with okadaic acid by anti-tumor promoters on BALB/3T3 cells
Acknowledgements
We thank Dr. T. Nishi and Otsuka Pharmaceutical Co. Ltd., for carrying out biological tests. We are
also grateful to Dr. M. Tanaka and Miss Y. Okamoto, Tokushima Bunri University, for measurements of
600 MHz NMR and MS spectra, respectively.

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