title: Enzyme-assisted extraction of anticoagulant polysaccharide from Liparis tessellatus eggs.
abstract: This study aimed to recover a heparin-like anticoagulant polysaccharide from Liparis tessellatus eggs (PLE) by using enzyme-assisted extraction technique. Extraction experiments were carried out using three different enzymes (Alcalase®2.4 L, Flavourzyme®500 MG, and Protamex®) under different conditions of temperature (45, 50, and 55°C), pH (6.5, 7.0, and 7.5), incubation time (24, 36, and 48 h), and enzyme to substrate ratio (E/S=0.5, 1.0, and 1.5%, w/w), which were combined according to a D-optimal design. Statistical analysis of extraction results allowed identifying the variables with greater influence on the extraction yield, and selecting the conditions that maximize the PLE extraction. The best extraction results were achieved when using the Protamex® enzyme in an E/S ratio of 1.34% (w/w), pH 6.60, 47.40°C, during 26.50 h. Under these conditions, a polysaccharide yield of 2.10% (w/w) was obtained. Clotting time measurements, activated partial thromboplastin time, and prothrombin time for evaluation of the anticoagulant properties of PLE were determined and showed increasing activities in correlation with the concentrations used. In the final step, the heparin-like nature of PLE was confirmed by digestion with heparinases I, II, and III, which showed ΔDiHS-0S, ΔDiHS-6S, ΔDiHS-diS1, and ΔDiHS-diS2 at compositions of 0.04, 0.03, 0.35, and 0.24 mol/g, respectively.
authors.affiliation: Department of Seafood Science and Technology, The Institute of Marine Industry, Gyeongsang National University, Tongyeong 650-160, Republic of Korea; Western Visayas College of Science and Technology, College of Arts and Sciences, Burgos St., La Paz, Iloilo City, Iloilo, Philippines. Electronic address: berna2423@gmail.com.
authors.forename: Bernadeth F
authors.initials: BF
authors.lastname: Ticar
authors.affiliation: Department of Seafood Science and Technology, The Institute of Marine Industry, Gyeongsang National University, Tongyeong 650-160, Republic of Korea; Faculty of Biology, Universitas Gadjah Mada, Yogyakarta 55281, Indonesia.
authors.forename: Zuliyati
authors.initials: Z
authors.lastname: Rohmah
authors.affiliation: Western Visayas College of Science and Technology, College of Arts and Sciences, Burgos St., La Paz, Iloilo City, Iloilo, Philippines.
authors.forename: Carmelo V
authors.initials: CV
authors.lastname: Ambut
authors.affiliation: Department of Seafood Science and Technology, The Institute of Marine Industry, Gyeongsang National University, Tongyeong 650-160, Republic of Korea.
authors.forename: Yeung-Joon
authors.initials: YJ
authors.lastname: Choi
authors.affiliation: Department of Biotechnology, Delft University of Technology, Julianalaan 67, 2628 BC Delft, The Netherlands.
authors.forename: Solange I
authors.initials: SI
authors.lastname: Mussatto
authors.affiliation: Department of Seafood Science and Technology, The Institute of Marine Industry, Gyeongsang National University, Tongyeong 650-160, Republic of Korea. Electronic address: bdchoi@gnu.ac.kr.
authors.forename: Byeong-Dae
authors.initials: BD
authors.lastname: Choi
mesh_terms: D000818:Animals; D000925:Anticoagulants / Q000737:chemistry* / Q000302:isolation & purification / Q000494:pharmacology; D005591:Chemical Fractionation / Q000379:methods; D002851:Chromatography, High Pressure Liquid; D004531:Eggs; D005399:Fishes; D011134:Polysaccharides / Q000737:chemistry* / Q000302:isolation & purification / Q000494:pharmacology
chemical_list: D000925:Anticoagulants; D011134:Polysaccharides
keywords: Anticoagulant activity; D-optimal design; Enzyme-assisted extraction; Liparis tessellatus eggs; Polysaccharide