Distribution, determination and enhancement of phlorotannins from brown seaweed Ecklonia cava

Abstract

Abstract In this study, brown seaweed were examine for their potent biological activities. Only brown seaweed contains marine algal polyphenol phlorotannins. Phlorotannins are plant secondary metabolites, formed by the polymerization of phloroglucinol (1, 3, 5-trihydroxybenzene). In brown seaweed, phlorotannins function as defense against herbivore, microbes and the harmful effect of ultra violate radiation and also act as structural compounds in cell wall hardening. Phlortannins have some potent biological activity such as antioxidant and anti-inflammatory activities, radical scavenging activity, anti-allergic activity, anti-plasmin activity, bactericidal activity, HIV-1 reverse transcriptase and protease inhibitory activity, acetyl cholinesterase inhibitory activity and tyrosinase inhibitory activity.

The second part of this study was focused on the distribution of phlorotannins in the brown alga Ecklonia cava and also compare the efficiency of different pretreatment condition for phlorotannins extraction. Two major phlorotannins dieckol and phlorofucofuroeckol-A were isolated from E. cava. For quantification of the dieckol and phlorofucofuroeckol-A, Reverse Phase High Performance Liquid Chromatography (RP-HPLC) was used. HPLC method showed good accuracy (92.64 and 94.02%, respectively), precision (3.92 and 3.94%, respectively), and linearity (r> 0.996). Mature thalli contained 1.5-fold higher amount of dieckol (1.82 mg/g-dry tissue) than young thalli. Distribution of phlorotannins in tissues of E. cava showed that blade tissue contained more phlorotannins than stipe and holdfast. Among differently dried thalli, shadow dried tissue showed that approximately 90% or more dieckol and phlorofucofuroeckol-A were extracted comparing to lyophilized tissue. In case of sun dried and oven dried thalli, approximately 60% of phlorotannins were extracted compared to lyophilized tissue. Washed thalli with fresh water, boiled thalli and steamed thalli were deteriorative for extraction of the compounds.

The third part of the study was focused on the development of a new technique for determination of two bioactive major phlorotannins dieckol and phlorofucofuroeckol-A from the hot water and solvent extracts of three brown seaweed E. cava, Ecklonia stolonifera and Eisenia bicyclis by RP-HPLC. To the best of our knowledge, so far no report has been found on the analytical method for simultaneous determination of solvent and water soluble phlorotannins from the brown seaweed by RP-HPLC. Comparing to solvent extraction, recovery yield of dieckol and phlorofucofuroeckol-A from hot water extract of E. stolonifera, the most, was approximately 90%. The antioxidant activity was observed from 200 ？g mL-1 water extract of E. bicyclis (91%), followed by E. stolonifera (90%) and E. cava (74%), respectively, in comparison with the standard butylated hydroxytoluene (BHT; 82%). IC50 of dieckol (14.2 ？M) and phlorofucofuroeckol-A (18.5 ？M) showed almost 9- and 7-fold stronger antioxidation than BHT (127.85 ？M). Therefore, the brown seaweed might be used as a potential source of phlorotannins, which can be utilized as a heat-stable, water-soluble, and antioxidant-rich functional food ingredient.

The fourth part of the study was focus on the effects of exogenous Methyl Jasmonate (MeJA) on the phlorotannin compounds of E. cava. E. cava was treated with various concentrations of MeJA (0 to 10 ？M) in PES media. Individual phlorotanin content was measured by RP-HPLC. Crude phlorotannins was increased by 56% compared to control. Highest amounts of individual phlorotannins such as dieckol, phlorofucofuroeckol-A, phlorotannin 1 and 2 were also observed in 2 ？M MeJA treatment, which were 85%, 53%, 118% and 37%, respectively higher than the control. For the time course, highest amount of crude and individual phlorotannins were significantly increased after 24 hrs of MeJA treatment. MeJA treatment also significantly increased the viability of E. cava tissue. These results indicate that MeJA can effectively enhance the phlorotannins compounds and also reduce the post harvest decay in E. cava.