Hydrolysis of sucrose solution into glucose and fructose by common ion-exchange resin (IR 118, IR 120, C 100 and C 120 E) in tall column was investigated. All resins in H(+)-form showed a better hydrolysis capability than their Ca(++)-Form. IR 118 (H(+)-Form) possessed the best hydrolysis property in comparison to other at the same flow rate and bed volumn. There was no mannose detected in chromatographic eluted solution, so there was no epimerization reaction occured in column during hydrolysis. The standard sugar mixture (sucrose, glucose and fructose) were then separated by IR 118 at 30 degree c and by PCR 833 (Purolite Chromatographic Resin) at 30, 40 and 60 degree c and it was found that separation at the rate of 60 ml/hr at 30 degree c using PCR 833 was the best condition for separating fructose given the output over 80%. PCR 833 was used for the separation of fructose out of the other invert sugar in sugar cane juice after hydrolysed with 1.5 % HCl at 70 degree c for 2 hours. The hydrolysate is composed of sucrose, glucose and fructose in the ratio of 1.5 : 8: 8. Consequently the best condition is the rate of 60 ml/hr at 30 degree c bed volume 720 ml. Decreasing the temperature from 60 degree c to 30 degree c will increase the efficiency of fructose separation. At the same time increasing the flow rate from 40 to 60 ml/hr will increase the fructose yield and the efficiency of separation increases when the volume of the resin is decreased.