Bali Medical Journal (Bali Med. J.) 2014, Volume 3, Number 3: 129-132 P-ISSN.2089-1180, E-ISSN.2302-2914 www.balimedicaljournal.org or www.ojs. unud.ac.id 129 THE ADDITION OF CAFFEINE IN EARLE’S BALANCED SALT SOLUTION MEDIA WITH WASHING UP METHOD INCREASE MEMBRANE INTEGRITY AND ACROSOMAL SPERM Bagus Komang Satriyasa and Agung Nova Mahendra Departement of Pharmacology, Faculty of Medicine, Udayana University, Bali-Indonesia Background: caffeine, a methylxanthine derivate, appears to inhibit phosphodiesterase, thereby inhibiting the break down of cAMP and increasing its concentration inside cell. This study aims to assess the effect of caffeine addition in Earles’s Balanced Salt Solution (EBSS) on the increase in membrane integrity and acrosome reaction of spermatozoa using swim up method. Methods: This study was carried out at the Clinic of Sexology and Andrology, Sanglah Public Hospital at Denpasar Bali-Indonesia. This study was an experimental study using the design of pre and post test paired control group design. The samples were sperm specimens of eighteen infertile couple male or volunteers who were infertile with age ranged between 20-40 years old. The samples were divided into two groups: treatment group (caffeine + EBSS) and control group (EBSS). The data were analysed statistically by normality test (Kolmogorov - Smirnov Goodness of Fit Test), Homogeneity test, and Paired Student’s t test. Results: The results showed that the caffeine addition in EBSS medium could increase significantly (p<0.05). The integrity of the sperm membrane obtained were from 81.30 % to 86.60 % and acrosomal reaction from 82.60% to 89.60% evaluated by hypo-osmotic swelling test (HOS). The conclusion of this study is that addition of caffeine in EBSS medium increases significantly membrane integrity and acrosomal reaction of the human sperm. Keywords: EBSS; caffeine; membrane integrity; and acrosomal reaction of sperm INTRODUCTION Sperm preparation should be carried out before aplying the Assisted Reproductive Technology (ART). 1,2 Some commonly known sperm preparation methods in Indonesia are swim up washing method (SUM), density gradient centrifugation (DGC), and side migration technique (SMT) (TMS). SUM technique is a favourite technique elaborated in many andrology clinics and the most commonly used medium in this process is Earli’s Balance Salt Solution (EBSS). In ART, pregnancy rate is relatively low reaching only 5% per cycle in IUI and 8% in IVF. 2 The cause of low achievement is multifactorial in nature. Some of the causative factors are the failure of spermatozoa to adhere onto the ovarial zona ifferen and the failure of acrosomal reaction to occur. The poor binding of ifference to ovum is commonly caused by defect in ifference membrane. 2,3 Substances or drugs is needed to enhance ifference membrane Address of Correspondence: Bagus Komang Satriyasa Departement of Pharmacology, Faculty of Medicine, Udayana University Bali-Indonesia Email: bsatriyasa@gmail.com integrity and acrosomal reaction, which ultimately lead to improved ifference property of the difference. One of them is caffeine (1,3,7-trimethyl- 2, 6-dioxy-purine). Caffeine, a phosphodiesterase inhibitor (PDEI) boosts ifference metabolic activity. Caffeine increases ifference motility, membrane integrity, and acrosomal reaction rate in vitro. 4,5 Khalili et al (1999) reported that membrane integrity of caffeine-washed spermatozoa is higher than those washed using standard medium (33,33% versus 27%). 6 Moreover, Matas et al (2002) reported that hypoosmotic swelling (HOS) test showed significant difference (p < 0,01) between sodium benzoate caffeine-treated spermatozoa and the control. 7 In this study, SUM technique was apllied, in order to study the effect of caffeine addition into EBSS media on the ifference membrane integrity and acrosomal reaction. The hypothesis of this study was the caffeine addition into EBSS media increases the membrane integrity and acrosomal reaction of SUM-harvested spermatozoa. RESEARCH METHODS The study was an experimental study with Pre-and post-test paired control group design. The sample counting was independent with average