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ABSTRACT: The culture conditions for arachidonic acid (AA)production by Mortierella alpina 1S-4 were investigated bymeans of a morphological study with the aims of obtaining ahigh AA yield and scale-up. In a 50-L jar fermentor study, amedium containing 3.1% soy flour and 1.8% glucose with 0.3%KH 2 PO 4 , 0.1% Na 2 SO 4 , 0.05% CaCl 2 ⋅2H 2 O and 0.05%MgCl 2 ⋅6H 2 O was found to be optimum. The AA yield reached9.8 g/L/7 d, and the major morphology was small pellets (1–2mm). However, in the case of the only addition of KH 2 PO 4 , themajor morphology was filaments. The apparent viscosity in-creased to 2240 cp, thereby requiring a high agitation speed tomaintain adequate oxygen tension, which caused mycelialdamage due to shear stress and therefore a decrease in the AAyield. When a medium with Na 2 SO 4 , CaCl 2 , and MgCl 2 wasused, the major morphology was large pellets (2–3 mm), andmass transfer limitation through the pellet wall caused a de-crease in the AA yield. Based on these results, a scale-up studywas carried out under the optimal conditions described above.An AA yield of 10.9 g/L/8 d was obtained in a 10-kL industrialfermentor, and the major morphology was small pellets.JAOCS 75, 1815–1819 (1998).KEY WORDS: Arachidonic acid, arachidonic acid yield, mineraladdition, morphology, Mortierella alpina, scale-up, viscosity. Arachidonic acid (5,8,11,14-cis-eicosapentaenoic acid, AA) hasvarious regulatory effects and physiological functions in hu-mans, and plays important roles in infant nutrition (1,2). Theimportance of AA has become more obvious recently, and therehave been some reports on AA production by Mortierella fungi(3–7) and safety evaluation of these fungal products (8,9). In theprevious study (10), we reported the effectiveness of mineraladdition to the basal medium; it was found to enhance the AAyield and to change the growth morphology.In general, sufficient mass transfer and oxygen supply arerequired to obtain high productivity in an aerobic fermenta-tion. In the case of a fungal fermentation, the medium com-position not only influences the productivity, but, as a side ef-fect, may also induce a change in growth morphology (11).The growth morphology has a strong effect on the physicalproperties of the fermentation broth, which causes numerousproblems in large fermentors with respect to gas dispersion,mass and heat transfer, and homogenization. Thus the fungalmorphology is often considered to be one of the key parame-ters in industrial fermentation. For the production of fungalmetabolic products, the desired morphology varies from oneproduct to another (12). Therefore, it is important to find theoptimal morphology for production and to study the influenceof minerals on the morphology.In spite of this obvious need, there have been few reportsof morphological studies on AA production by Mortierellafungi. For commercial production, a high biomass concentra-tion is essential because it is an intracellular product. Besides,AA production requires adequate oxygen, because AA isformed through enzymatic desaturation which comprises oxy-genation (13). Thus an adequate oxygen supply by means ofagitation, aeration, and morphological control is a key factorin obtaining a higher AA content in the cells. Therefore, weexamined the effects of mineral addition on the morphology,AA productivity and cell growth, and then attempted toachieve scale-up to an industrial fermentor by controlling theoptimal morphology.MATERIALS AND METHODSMicroorganism and culture conditions. Mortierella alpina 1S-4(3) was used throughout this study. A stock culture was storedon a Czapek-medium slant of 0.2% NaNO 3 , 0.1% K 2 HPO 4 ,0.05% MgSO 4 ·7H 2 O, 0.05% KCl, 0.001% FeSO 4 ⋅7H 2 O, 3%sucrose, and 2% agar, slant in a test tube at 5°C. For the jar fermentor study, an inoculum was prepared in a500-mL flask containing 100 mL medium including 1.8%glucose and 1% yeast extract, with shaking for 3 d at 28°C,and then the main culture was carried out in a 50-L jar fer-mentor (Mitsuwa Bio Systems Co., Ltd., Osaka, Japan) witha working volume of 25 L under such conditions as 26°C, aninoculation rate of 0.5%, an aeration rate of 25 L/min, and aheadspace pressure of 200 kPa. The agitation speed wasCopyright © 1998 by AOCS Press 1815 JAOCS, Vol. 75, no. 12 (1998)*To whom correspondence should be addressed.E-mail: Kenichi_Higashiyama@suntory.co.jpEffects of Mineral Addition on the Growth Morphologyof and Arachidonic Acid Productionby Mortierella alpina 1S-4Kenichi Higashiyama a,* , Toshiaki Yaguchi a , Kengo Akimoto a ,Shigeaki Fujikawaa, and Sakayu Shimizu ba Institute for Fundamental Research, Suntory Limited, Yamazaki 5-2-5, Shimamoto-cho, Mishima-gun, Osaka 618-0001,Japan, and b Department of Agricultural Chemistry, Kyoto University, Sakyo-ku, Kyoto 606-8224, Japan