J.Exp. Life Sci. Vol. 10 No. 2, 2020 ISSN. 2087-2852 E-ISSN. 2338-1655 Cold Storage Time on Malondialdehyde (MDA) Level, Motility and Viability Spermatozoa (Wirastuti, et al.) 127 Effect of Cold Storage Time (4°C) on Malondialdehyde (MDA) Level, Motility and Viability Spermatozoa of Cyprinus carpio L. Punten Strain Rosyi Wirastuti 1 , Agung Pramana Warih Marhendra 2 , Jantje Wiliem Souhaly 2 , Sri Rahayu 2* 1 Master Program of Biology, Faculty of Mathematic and Natural Sciences, University of Brawijaya, Malang, Indonesia 2 Department of Biology, Faculty of Mathematics and Natural Sciences, University of Brawijaya, Malang, Indonesia Abstract The aim of this research is to know the level of MDA, motility, and viability of spermatozoa of Cyprinus carpio L. in cold storage time at 4°C. This study used treatment extenders that were NaCl 0.9% as control, diluent of egg yolk with concentrate 0%, 5%, 10%, 15%, and storage in 0 h, 24 h, 48 h, 72 h, and 96 h at 4°C. The data were analyzed using ANOVA (P<0.05). The results showed that the quality (motility and viability) of spermatozoa C. carpio L. Punten strain in cold storage could be maintained at 96 h. The optimum storage for motility was found in a diluent of egg yolk concentrate 5% at 48 h. MDA levels of semen from C. carpio L. Punten strains after being stored at cold temperatures increased at 0 h to 24 h, then decreased after 48 h of storage and increase at 72 h - 96 h stored. Egg yolk concentration and storage time had a significant effect on MDA levels. There is a negative correlation between MDA levels and motility. There was no correlation between MDA levels and the viability of spermatozoa C. carpio L. Punten strain. Keywords: Cyprinus carpio L, Egg yolk, Malondialdehyde, Motiliy, Viability. INTRODUCTION  Cyprinus carpio L is classified as a type of fish that has high economic value [1]. One of the main factors needed to increase the cultivation and production of C. carpio L is created as the best quality of parental fish to produce good quality seeds. The process of producing the best quality of parental fish required a long time and high cost. Therefore, it must be utilized optimally. During the reproductive season, parental fish sperm can be stored and can be used when required. The advantage of sperm storage is can be stored for a long time and can be used when necessary [2]. Sperm storage at low temperatures can cause cold shock [3], which can produce ROS (Reactive Oxygen Species) [4]. ROS is highly reactive, resulting in lipid peroxidation. Malondyladehid (MDA) is the final result of the lipid peroxidation process, which is toxic to cells. MDA can cause damage and decrease the integrity of the spermatozoa membrane, resulting in decreased sperm quality [5]. Sperm storage required diluent which protects sperm from low temperatures and provides an energy source during the storage process, and also to keep sperm from damaged and die during storage. The quality of sperm during storage is maintained using a good medium. The use of Correspondence address: Sri Rahayu Email : srahayu@ub.ac.id Address : Faculty of Mathematic and Natural Science, Brawijaya University, Malang, 65145 storage media can inhibit sperm movement, stabilize the physiology of sperm, and maintain the viability of sperm [6]. The diluent of sperm was supply nutrients and maintained the osmotic pressure and electrolyte balance to protected viability sperm. One of the diluents that can be used is egg yolk. Since 60 years ago, egg yolk has been shown to be an effective medium for storing sperm in various species, for example, the storage of salmon (Salmon salar) and rainbow trout (Oncorhynchus mykiss) [7]. Egg yolk contains LDL (Low-Density Lipoprotein), which can maintain sperm at low temperature, increase motility during storage, and binding to the sperm membrane to preserving the membrane from protein damage [8]. The content of egg yolks such as cholesterol, fatty acids, phospholipids, progesterone, and antioxidant compounds have the quality as a protective agent [9,10,11]. Therefore, it is necessary to choose a suitable and efficient medium storage of egg yolk. The research aimed to determine the quality and spermatozoa MDA levels of C. carpio L. Punten strain after cold stored at temperature 4°C with diluent of egg yolk, and evaluate the interaction between MDA levels with motility and viability of spermatozoa. MATERIAL AND METHOD Semen Collection Semen was collected from six male C. carpio L. Punten strain aged one year and the weight ± 700 g by manual abdominal stripping.