Journal of Molecular Catalysis B: Enzymatic 56 (2009) 7–12 Contents lists available at ScienceDirect Journal of Molecular Catalysis B: Enzymatic journal homepage: www.elsevier.com/locate/molcatb Optimized synthesis of lipase-catalyzed l-ascorbyl laurate by Novozym ® 435 S.-W. Chang a , C.-J. Yang a , F.-Y. Chen a , C.C. Akoh b , C.-J. Shieh a,c,∗ a Department of Bioindustry Technology, Dayeh University, 112 Shan-Jiau Road, Da-Tsuen, Chang-Hua 515, Taiwan b Department of Food Science and Technology, University of Georgia, Athens, GA 30602, USA c Biotechnology Center, National Chung Hsing University, Taichung 402, Taiwan article info Article history: Received 31 October 2007 Received in revised form 7 April 2008 Accepted 7 April 2008 Available online 16 April 2008 Keywords: Antioxidant Ascorbyl esters Lipase Optimization Response surface methodology abstract l-Ascorbyl laurate is a fatty acid derivative of l-ascorbic acid which can be widely used as a natural antioxidant in both lipid containing food and cosmetic applications. To avoid any possible harmful effects from chemically synthesized product, the enzymatic synthesis appears to be the best way to satisfy the consumer demand for natural antioxidants. The ability of immobilized lipase from Candida antarctica (Novozym ® 435) to catalyze the direct esterification between l-ascorbic acid and lauric acid was inves- tigated. Response surface methodology (RSM) and 5-level-4-factor central composite rotatable design (CCRD) were employed to evaluate the effects of synthesis parameters, such as reaction time (2–10h), temperature (25–65 ◦ C), enzyme amount (10–50% w/w of l-ascorbic acid), and substrate molar ratio of l-ascorbic acid to lauric acid (1:1–1:5) on percentage molar conversion to l-ascorbyl laurate. Based on the analysis result of ridge max, the optimal enzymatic synthesis conditions were predicted as follows: reac- tion time 6.7 h, temperature 30.6 ◦ C, enzyme amount 34.5%, substrate molar ratio 1:4.3; and the optimal actual yield was 93.2%. © 2008 Elsevier B.V. All rights reserved. 1. Introduction Polyphenols are useful antioxidants that can protect cells or DNA from damage by free radicals and are widely used by the food and cosmetic industries. Among the available antioxidants, butylated hydroxyanisole (BHA) and butylated hydroxytoluene (BHT) are the most common polyphenols used. However, they might possibly generate toxic or carcinogenic components as they degrade during storage [1]. l-Ascorbic acid (vitamin C), a natural hydrophilic antioxidant, has been used but limited in its applica- tion in hydrophobic foods and cosmetics [2]. To overcome solubility problem, a fatty acid derivative of l-ascorbic acid, such as l-ascorbyl laurate which possesses similar antioxidant function as l-ascorbic acid was used to prevent the enzymatic browning of apple juices or mixed with other fatty acid esters of ascorbic acid for treating and preventing sunburn damage to the skin [3,4]. Traditionally, such polyphenol compounds could be synthesized by chemical meth- ods. Due to the steady growing demand for natural materials, the biosynthesis of such esters by lipase-catalyzed reactions, under mild conditions, has become a current commercial interest. An opti- ∗ Corresponding author at: Biotechnology Center, National Chung Hsing Univer- sity, Taichung 402, Taiwan. Tel.: +886 4 2284 0452x5121; fax: +886 4 2285 2609. E-mail address: cjshieh@nchu.edu.tw (C.-J. Shieh). mized enzymatic synthesis of l-ascorbyl esters with improved yield at reduced cost in the most favorable conditions would be more appealing to the consumer and of benefit to the manufacturers. The importance of the enzymatic synthesis, catalyzed by lipases, to produce l-ascorbyl esters via esterification in water-miscible organic solvents has been emphasized in several works [1,5,6]. The lipase-catalyzed esterification reactions were reviewed, including the parameters affecting the lipase activities on esterification reac- tions such as reaction time, synthesis temperature, added water, water activity, pH memory, and acyl donors. [7]. Response surface methodology (RSM) and central composite rotatable design (CCRD) are useful statistical techniques for complex processes investigation and have been successfully applied to optimizing ester production by lipase [8]. Humeau et al. [1] synthesized ascorbyl palmitate through trans- esterification by using an immobilized lipase (Novozym ® 435) from Candida antarctica in nonaqueous environment containing a 1:5 ini- tial molar ratio of ascorbic acid to acyl donor. When palmitic acid methyl ester was used as acyl donor, 68% of ascorbic acid was con- verted to the ascorbyl ester compared to 56% with palmitic acid. Humeau et al. [1] also reported the enzymatic synthesis of fatty acid ascorbyl esters by lipase B (C. antarctica) in 2-methyl-2-butanol. They utilized palmitic acid methyl ester, EPA ethyl ester and DHA ethyl ester as acyl donors to react with l-ascorbic acid in the transes- terification reactions. When methyl palmitate/ascorbic acid molar 1381-1177/$ – see front matter © 2008 Elsevier B.V. All rights reserved. doi:10.1016/j.molcatb.2008.04.001