Synthesis of novel phospholipids that bind phenylalkanols and hydroquinone via phospholipase D-catalyzed transphosphatidylation

Abstract

Phenylalkanols such as tyrosol and hydroxytyrosol (h-tyrosol), which possess antioxidant and anticancer properties, were phosphatidylated by phospholipase D (PLD)-catalyzed transphosphatidylation. After a 24-h reaction of phosphatidylcholine (PC) and tyrosol with PLD, a new product was detected by TLC and identified to phosphatidyl-tyrosol by high resolution MS and NMR analyses. The optimum reaction conditions were as follows; soyPC 50 µmol, tyrosol 500 µmol, ethyl acetate 1.6 ml, PLD 1.6 U, 0.2 M sodium acetate buffer (pH 5.6) 0.8 ml, 37 o C for 24 h. Under the optimum reaction conditions, the yields of phosphatidyl-tyrosol, hydroquinone (HQ), 2-(4-aminophenyl)ethanol (4APE), h-tyrosol, and 2-phenylethanol (PEA) were 87±3.7, 13±1.3, 90±2.3, 64±5.5, and 85±1.0 mol%, respectively. Furthermore, from the results of transphosphatidylation of soyPC with several phenylethanols and phenylpropanols, we established the following details about the reaction specificity of transphosphatidylation by PLD from Streptomyces sp: (1) Para-amino and para-hydroxyl groups in the benzene ring of PEA derivatives do not affect the transphosphatidylation by PLD, whereas meta-hydroxyl group slightly inhibits the transphosphatidylation. (2) Meta and ortho-methyl groups in the benzene ring of PEA derivatives also slightly inhibit the transphosphatidylation. (3) Secondary and tertiary alcohols and hydroquinone are difficult to transphosphatidylate by PLD. 3 IntroductionPhospholipase D (PLD) (EC 3.1.4.4) is a lipolytic enzyme used for hydrolyzing the terminal phosphodiester bond of phospholipids (PLs) [1]. PLD can also be used as a catalyst in the transphosphatidylation in order to exchange the phosphatidyl moiety of PLs to various alcohols.Transphosphatidylation is an effective reaction for the application of functional alcohols in a wide variety of fields such as the production of fine chemicals, functional foods. In previous studies, 6-phosphatidyl-L-ascorbic acid was synthesized from egg yolk phosphatidylcholine (PC) and L-ascorbic acid [2], it exhibited a high anti-oxidative activity [3]. In addition, 5-fluorouridine[4], kojic acid and arbutin [5] were phosphatidylated by PLD-catalyzed transphosphatidylation, and the synthesized compounds exhibited antitumor effects [4] and inhibitory effects on tyrosinase [5]. We have also reported the synthesis of phosphatidyl-terpenes such as phosphatidyl-geraniol, phosphatidyl-myrtenol, and phosphatidyl-perillyl alcohol by PLD [6,7].The synthesized phosphatidyl-perillyl alcohol significantly reduced viability of human leukemia and prostate cancer cells [7].In this study, we focused on the transphosphatidylation of functional phenylalkanols by PLD.2-(4-Hydroxyphenyl)ethanol (tyrosol; 1) and 2-(3,4-dihydroxyphenyl)ethanol (h-tyrosol; 5) ( that has an anxiolytic-like effect on mice [11], and tyrosinase inhibitory activity [12]. In addition, we used hydroquinone (HQ; 3), 2-(4-aminophenyl)ethanol (4APE; 4), 2-phenylethanol (PEA; 6) ( Fig. 1) and other phenylalkanols as substrates to clarify the reactio...

Phospholipase D (PLD) catalyzed synthesis of phosphatidyl-glucose in biphasic reaction system

“…Thus, an excess of glucose will theoretically shift the reaction equilibrium towards transphosphatidylation and formation of PL-Glu. Nevertheless, an overly excessive amount of acceptor alcohol was reported to inhibit the transphosphatidylation activity of PLD (Yamamoto et al, 2008(Yamamoto et al, , 2011. In addition, excess of glucose was also reported to inhibit the esterification activity of lipase by distorting the enzyme-water interaction and even stripped water from enzyme (Moreau et al, 2007).…”

“…Early studies on enzymatic transphosphatidylation involved the modification of a choline head group in lecithin with serine, glycerol or ethanolamine for the production of the respective PLs. Meanwhile, the current focus in enzymatic transphosphatidylation is to produce novel structured PLs such as phosphatidyl-terpenes (Yamamoto, Hosokawa, Kurihara, & Miyashita, 2008), phosphatidyl-sterol (Hossen & Hernandez, 2004), phosphatidyl-phenylalkanol (Yamamoto, Kurihara, Miyashita, & Hosokawa, 2011), phosphatidyl-Tris, phosphatidyl-diethylamine/triethylamine, and phosphatidyl-BisTris (Dippe, Mrestani-Klaus, Schierhorn, & Ulbrich-Hofmann, 2008). These novel structured PLs are usually designed with improved functionalities such as anticancer or antioxidant (Yamamoto et al, 2008(Yamamoto et al, , 2011 properties.…”

Enzymatic synthesis and modification of structured phospholipids: recent advances in enzyme preparation and biocatalytic processes

“…Compared with aliphatic alcohols, there is little information on the synthesis of PLs using aromatic alcohols. Recently, functional phenylalkanols such as tyrosol and hydroxytyrosol, which exhibit antioxidant and anticancer properties, have also been introduced to PLs by PLD-catalyzed transphosphatidylation (Yamamoto et al 2011). On the basis of the results, the details about the reaction specificity of PLD from a Streptomyces sp.…”

Novel glyceryl ethers phospholipids produced by solid to solid transphosphatidylation in the presence of a food grade phospholipase D

“…Enzymatic transphosphatidylations usually reported in the literature, utilize very low concentration of PL and a large excess of the other reactant . In general, these processes are performed in a biphasic stirred emulsion systems, comprised of an aqueous buffer and an organic solvent.…”