Fertilization is one of the most important procedures in artificial reproduction and it directly affects the reproduction outcome. When there is optimization of fertilization, there can be a positive effect on subsequent reproductive processes and economic aspects of aquaculture. This study was conducted to determine time for which oocytes and sperm of barbel Barbus barbus retain fertilization capacity following placement in freshwater. Furthermore, the amount of ovarian fluid, excreted by fish during spawning with eggs (OFI; %) was determined, along with the chemical composition and effects on fertilization were determined. Gametes, ovarian fluid, and seminal plasma from barbel spawning specimens of the F4 generation were used to conduct the study. Ovarian fluid accounted for 14%-68% of contents of the mass released at spawning and post-spawning composition differed depending on whether hormonal treatments were utilized for control of reproduction. There was an association (R2 = 0.982; P = 0.000) between the pH of ovarian fluid and the barbel embryo survival rate. There was the greatest survival rate (>60 %) when the pH range of 7.9-8.4 and there was a lesser embryo viability when pH values were lesser or greater than values within this range (P < 0.05). The results from the study indicate that barbel eggs retain fertilization capacity longer (as long as 210 s) after activation by placement in fresh water than spermatozoa (about 30 s). Copyright © 2020 The Author(s). Published by Elsevier B.V. All rights reserved.