Extraction of natural dye from the roots of Rubia Cordifolia by conventional extraction, as well as by using ultrasonic extraction, has been performed in this study. Process variables, namely ultrasonic frequency (25, 40, 58 kHz), solid-to-liquid ratio (1:5, 1:10, 1:15) and extraction time (2, 2.5, 3 h) were studied, and the optimum condition was determined using three-factor Central Composite Design (CCD). The optimum extraction condition obtained from CCD was 25 kHz ultrasonic frequency, 1:15 solid-to-liquid ratio and extraction time of 3 h. The experimental result obtained at the optimum condition is in good agreement with the predicted value. An Artificial Neural Network(ANN) model was also developed, and the predictions are compared with values obtained from Response Surface Methodology (RSM) model. The extracted compounds were identified using TLC and FT-IR Spectroscopy. The extracted dye was used for dyeing leather and cotton fabric along with different mordants to obtain different shades. (C) 2017 Elsevier B.V. All rights reserved.