International Journal of Biological Macromolecules 50 (2012) 624–631 Contents lists available at SciVerse ScienceDirect International Journal of Biological Macromolecules jo u rn al hom epa ge: www.elsevier.com/locate/ijbiomac Protective effects of aminoethyl-chitooligosaccharides against oxidative stress in mouse macrophage RAW 264.7 cells Dai-Nghiep Ngo a,b , Moon-Moo Kim c , Se-Kwon Kim a,d,∗ a Marine Bioprocess Research Center, Pukyong National University, Busan 608-737, South Korea b Department of Biochemistry, Faculty of Biology, University of Science, VNU-HCM, Ho Chi Minh City, Viet Nam c Department of Chemistry, Dong-Eui University, Busan 614-714, South Korea d Department of Chemistry, Pukyong National University, Busan 608-737, South Korea a r t i c l e i n f o Article history: Received 13 December 2011 Received in revised form 18 January 2012 Accepted 25 January 2012 Available online 4 February 2012 Keywords: Aminoethyl-chitooligosaccharides (AE-COS) Oxidative stress Free radical scavenging Antioxidant RAW 264.7 cells a b s t r a c t The aim of this study is to investigate the inhibitory effects of aminoethyl-chitooligosaccharides (AE-COS) on oxidative stress in mouse macrophages (RAW 264.7 cells). The inhibitory effects of AE-COS on DNA and protein oxidation were studied in RAW 264.7 cells. Furthermore, free radical scavenging effect of AE- COS were determined in RAW264.7 cells by 2 ′ ,7 ′ -dichlorofluorescein (DCF) intensity and intracellular glutathione (GSH) level. AE-COS also inhibited myeloperoxidase (MPO) activity in human myeloid cells (HL-60). These results suggest that AE-COS acts as a potential free radical scavenger in RAW 264.7 cells. © 2012 Elsevier B.V. All rights reserved. 1. Introduction In biological systems, equilibrium between oxidants formation and endogenous antioxidant defense mechanisms exists to pro- tect cellular biomolecules against oxidation. The oxidative stress will happen if that balance is disturbed [1]. Oxidative stress causes injury to important cellular components, thus the reac- tive oxygen species (ROS) generated excessively in tissues can lead to death of cells. Furthermore, ROS play an important role in many diseases such as cancer, arthritis, neurodegenerative, dia- betes, hypertension, inflammation and aging. They have direct or indirect relationship with oxidation of cellular biomolecules [2,3]. Therefore, the studies to develop novel antioxidant are necessary. Chitosan is prepared with deacetylation of chitin, a polysac- charide abundantly found in nature in the presence of alkali at high temperature. It has numerous biological activities such as immuno-enhancing activity, antitumor activity [4], antibacterial activity [5], antifungal activity [6], anti-hypertensive effects, and antioxidant activity [7–9]. Chitooligosaccharides (COS), partially ∗ Corresponding author at: Marine Bioprocess Research Center, Pukyong National University, Busan 608-737, South Korea. Tel.: +82 51 629 7094; fax: +82 51 629 7099. E-mail address: sknkim@pknu.ac.kr (S.-K. Kim). hydrolyzed products of chitosan, is of great interest in pharmaceu- tical and medicinal applications due to their non-cytotoxic and high water soluble properties [7]. Moreover, the structure and proper- ties of chitosan and its derivatives have been studied more and more. These reports helped to improve the structural properties of chitosan and the similar structural compounds for a particu- lar application by chemical modification. However, research on synthesis of COS derivatives and identification of their biological activities have been seldom reported. Therefore, our objective was to develop a COS derivative, aminoethyl-chitooligosaccharides (AE- COS), with improved antioxidant activity in mouse macrophage RAW 264.7 cells. 2. Materials and methods 2.1. Materials Chitooligosaccharides (MW 800–3000 Da, degree of deacety- lation, DD, 90%) prepared from crab shells were donated by Kitto Life Co. (Seoul, Korea). MTT reagent (3-(4,5-dimethyl-2-yl)- 2,5-diphenyltetrazolium bromide), 3,3 ′ ,5,5 ′ -tetramethybenzidine (TMB), 2,4-dinitro-phenyl hydrazine reagent, 2 ′ ,7 ′ - dichlorofluorescein diacetate (DCFH-DA), agarose, and fetal bovine serum (FBS) and some other chemicals were purchased from Sigma Chemical Co. (St. Louis, MO, USA). 0141-8130/$ – see front matter © 2012 Elsevier B.V. All rights reserved. doi:10.1016/j.ijbiomac.2012.01.036