Background:The utilization of Rhizopus oligosporus to produce fatty acid omega-3 in liquid substrate has been conducted widely. The content of fat in the palm oil waste in the amount of 5.56/100 gram still has potential to produce fatty acid omega-3. The solid fermentation of tofu waste with R. oligosporus could increase the content of fatty acid up to 34.4% and palm oil waste could increase up to 61.57%. Methods: Sample is collected from the industry of palm-oil. The palm-oil waste is considered as a substrate, and the mold used is R.oligosporus. The waste product is formulated by adding urea and sucrose. In this case, urea is used as the source of N (nitrogen) and sucrose is used as the source of C (carbon). The substrate with nitrogen is called as low-carbon treatment and the substrate with carbon is called as high-carbon treatment. For the controlle we did not add anything. Fermentation is conducted within 7 days on the shaker at room temperature. The product of fermentation was analyzed for the Water, Ash, Fat, and Fatty-Acid Omega-3 contents. Results: This research shows that the water content of the fermentation product decreased and ash content increased. This happened both in the low-carbon treatment and in the high-carbon treatment. The content of fat after fermentation in controlle treatment increased by 6.44%. However, in the low-carbon treatment fermentation, the fat content decreased by 31.14% from the original content. In the high-carbon treatment fermentation, the fat content increased by 31.67%. The content of saturated-fatty acids produced in the high-carbon treatment and low-carbon treatment are as follows: Capric acid (10:0) and lauric acid (12:0) decreased and myristic acid (14:0), palmitic acid (16:0) and stearic acid (18:0) increased, except the myristic-acid in high-carbon treatment decreased . The content of unsaturated-fatty acid in controlle and high-carbon treatment: Oleic acid (18:1), linoleic acid (18:2) and linolenic (18:3) increased. However, all fatty acid in low-carbon treatment decreased, except the linolenic-acid. The conclusion: The fermentation of palm-oil waste with Rhizopus oligosporus mold could increase the content of fat and produce fatty acid omega-3. In addition, the high-carbon substrat could increase the production of unsaturated-fatty acid. Submit : 19-12-2011 Review : 08-03-2012 Review : 12 -03-2012 revisi : 17–4-2012 56 Keywords: liquefied-fermentation, waste product of palm oil, R.oligosporus, fatty acid 0mega-3 Abstrak Latar belakang: Pemanfaatan kapang Rhizopus. oligosporus untuk menghasilkan asam lemak omega-3 pada substrat cair telah banyak dilakukan. Kandungan lemak ampas kelapa sawit 5,56 gram/100 gram masih berpotensi untuk menghasilkan asam lemak omega-3. Fermentasi padat pada substrat ampas tahu dan ampas kelapa sawit dengan kapang Rhizopus. oligosporus dapat meningkatkan kadar lemak: ampas tahu 34,4%, sedangkan pada substrat ampas kelapa sawit dengan formula tinggi karbon, kadar lemak meningkat 61,57%. Metoda: Sampel ampas sawit diambil dari pabrik industri minyak sawit. Pada penelitian ini ampas sawit dipakai sebagai substrat fermentasi dan kapang yang digunakan adalah R.oligosporus. Untuk bahan suplemen digunakan urea dan sukrosa Kontrol adalah ampas-sawit tanpa suplemen, sedangkan perlakuan ampas sawit ditambahkan urea sebagai sumber Nitrogen(N) dan ampas sawit ditambah sukrosa sebagai sumber Karbon(C). Penambahan sumber N sebagai substrat rendah karbon dan sumber C sebagai substrat tinggi karbon. Fermentasi dilakukan selama 7 hari diatas shaker pada suhu ruang. Produk hasil fermentasi dilakukan analisis: kadar air; abu, lemak, dan asam lemak omega-3. Hasil penelitian: Hasil menunjukkan bahwa kadar air produk hasil fermentasi menurun pada kontrol dan semua perlakuan. Kadar abu meningkat untuk semua perlakuan. Kandungan lemak pada ampas kontrol dan ampas-sukrosa meningkat 6,43% dan 31,67%, sedang substrat ampas-urea menurun 31,14%. Kandungan asam lemak jenuh; asam kaprat (10:0) dan laurat (12:0) menurun, sedangkan untuk asam miristat (14:0), palmitat (16:0), dan stearat (18:0) meningkat. Kandungan asam lemak tidak jenuh pada substrat ampas kontrol dan ampas-sukrosa: oleat (18:1), linoleat (18:2), dan linolenat (18:3) semua meningkat, sedangkan substrat ampas-urea untuk oleat (18:1) dan linolenat (18:3) menurun, kecuali linoleat (18:2) meningkat. Asam palmitoleat (16:1) tidak terdeteksi. Kesimpulan: Fermentasi cair ampas kelapa sawit dengan kapang Rhizopus oligosporus dapat meningkatkan kandungan lemak dan menghasilkan asam lemak omega-3. Selain itu, substrat tinggi karbon dapat meningkatkan produksi asam lemak tidak jenuh. Kata kunci: fermentasi cair, ampas sawit, R oligosporus, asam lemak omega-3