Elsevier

Fitoterapia

Volume 80, Issue 4, June 2009, Pages 233-236
Fitoterapia

Regioselective acylation of 3-O-angeloylingenol by Candida antarctica Lipase B

https://doi.org/10.1016/j.fitote.2009.02.001Get rights and content

Abstract

Acylation of 3-O-angeloylingenol (1) with vinyl acetate, vinyl decanoate and vinyl cinnamate, catalyzed by Candida antarctica Lipase B, was investigated. In each case, compound 1 was quantitatively and regioselectively acylated to afford a single product, 3-O-angeloyl-20-O-acetylingenol (1a), 3-O-angeloyl-20-O-decanoylingenol (1b) and 3-O-angeloyl-20-O-cinnamoylingenol (1c), respectively. The structures of the novel compounds 1b1c were determined by MS and NMR, and product 1a by comparison of RP-HPLC and TLC with a standard. Compounds 1b1c induced a bipolar morphology of MM96L melanoma cells at a similar concentration as compound 1, as well as having activity in inhibiting the growth of MM96L melanoma cells.

Introduction

Candida antarctica Lipase B (commercial name: Novozyme 435) is a lipase which has proven to be a highly regio- and stereo-selective enzyme in both hydrolysis [1] and synthesis [2]. This enzyme has been used to acylate a variety of substrates, including many classes of natural products [3]. We successfully used C. antarctica Lipase B for the regio- and stereo-selective acylation of ginsenosides [4], iridoid glycosides, a flavanoid glycoside and simple phenolic compounds [5].

3-O-angeloylingenol (1) was isolated from Euphorbia antiquorum and E. peplus, the extracts of which have been used in the treatment of a number of conditions, such as warts, corns, waxy growth and skin cancer [6]. First characterized as a skin irritant and a toxic Euphorbia factor, An1 [7], 3-O-angeloylingenol (1) was thereafter shown to induce in vitro apoptosis of melanoma cells [8] and leukemia cells [9]. This compound showed potential as a novel topical chemotherapeutic agent for the treatment of skin cancer [10] and is being developed as a topical therapy for actinic keratosis and superficial basal cell carcinoma (www.peplin.com).

In this study, 3-O-angeloylingenol (1) was acylated with vinyl acetate, vinyl decanoate and vinyl cinnamate in the presence of C. antarctica Lipase B. The acylated products 1a1c were isolated, purified and their structures determined by MS and NMR (Fig. 1). The activity of compounds 1b1c in inhibiting the growth and inducing a bipolar morphology of MM96L melanoma cells was assessed.

Section snippets

General

Vinyl acetate, vinyl decanoate, vinyl cinnamate and 2-methylbutan-2-ol were purchased from Sigma. C. antarctica Lipase B (commercial name: Novozyme 435), immobilized on acrylic resin, was a kind gift from Novozymes Australia Pty Ltd. RPMI 1640 media were purchased from Life Technologies, USA, fetal calf serum and 96-well tissue culture plates from CSL Biosciences, Australia.

Standards of 3-O-angeloylingenol and 3-O-angeloyl-20-O-acetylingenol were from Peplin Ltd., Australia. Pre-coated thin

Results and discussion

3-O-angeloylingenol (1) was reacted with either vinyl acetate, vinyl decanoate or vinyl cinnamate in the presence of C. antarctica Lipase B. The reactions were monitored by TLC and HPLC. Small scale experiments indicated that 3-O-angeloylingenol (1) was quantitatively acylated into a single product in the presence of each of the three acyl donors. To generate sufficient amounts of products for structural determination, the reactions were scaled-up and the acylated products were then purified by

Acknowledgements

This work was funded by a grant from the Australian Government to the CRC for Bioproducts. We wish to thank Novozymes Australia Pty Ltd. for providing Novozyme 435, and Drs. T. K. Lim and Frances Separovic, School of Chemistry, The University of Melbourne, Australia, for access to the NMR spectrometer. We also thank Dr Ming-Long Liao, CRC for Bioproducts, The University of Melbourne, Australia, for critical comments.

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Current address: Nutrifina Pty Ltd., 103 Albert St, Port Melbourne, Victoria 3207, Australia.

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