Acanthopanax species is known commonly as Siberian ginseng, touch-me-not, devil’s shrub, prickly eleutherococc, eleutherococc and wild pepper. A diverse group of chemical compounds isolated from Acanthopanax species was named ‘eleutherosides’. Among eleutherosides, eleutherosides B and E were widely known in Acanthopanax species. Acanthopanax species are cultivated and grow wild in a various area of Korea and have a variety of pharmacological effects. But, there are a lot of difficulties on producing excellent Acanthopanax species, according to the cultivated method is different pharmacological ingredients. This study, therefore, analyzed eleutherosides B and E in A. divaricatus and A. koreanum by different fertilizer ratio using HPLC. We will be investigated a high content of eleutherosides B and E by different fertilizer ratio and suggest an efficient fertilizer ratio of A. divaticatus and A. koreanum. All samples of A. divaricatus and A. koreanum were collected at Yeongcheon Agricultural Technology & Extension Center, Yeongcheon, Korea. The sample was prepared by upper and lower parts. The fertilizer ratio are N-P-K(10.5-8.5-8.5: 50 kg/10a), 2N-P-K (21-8.5-8.5: 50 kg/10a), N-2P-K (10.5-17-8.5: 50 kg/10a), N-P-2K (10.5-8.5-17: 50 kg/10a), and 2N-2P-2K (21-17-17: 50 kg/10a), respectively. To analyze eleutherosides B and E, 5 g of A. divaricatus and A. koreanum was extracted with 50% MeOH (3 × 100 ml) by reflux and evaporated in vacuo. The residue was dissolved in 1 ml of MeOH. The resulting solution was used for HPLC analysis. HPLC separation of eleutherosides B and E for qualitative and quantitative analysis was performed using a reverse phase system. A Discovery®C18 (4.6 × 250 mm, 5 μm) column was used with a mobile phase that consisted of water and acetonitrile. A gradient solvent system of water and acetonitrile (90:10 to 70:30 for 20 min) was used for the elution program. UV detection was conducted at 350 nm. The injection volume was 10 μl and the flow rate was 1 ml/min. All injections were performed in triplicate. The different fertilizer ratio yielded total eleutherosides B and E contents of 4.417-6.905 and 3.652-7.227 mg/g in the upper and lower parts of A. divaricatus, respectively. In A. koreanum, the total eleutherosides B and E contents were 4.591-10.108 and 3.834-9.079 mg/g in the upper and lower parts, respectively. The best conditions to increase eleutherosides B and E content in A. divaricatus was determined to be with N-2P-K fertilizer ratio, on the other hand, in A. koreanum was 2N-2P-2K fertilizer ratio.