Biotechnol. Appl. Biochem. (2003) 38, 193–199 (Printed in Great Britain) 193 Optimization of submerged-culture conditions for mycelial growth and exo-biopolymer production by Auricularia polytricha (wood ears fungus) using the methods of uniform design and regression analysis Chun-Ping Xu and Jong-Won Yun 1 Department of Biotechnology, Daegu University, Kyungsan, Kyungbuk 712-714, Korea This paper is concerned with the optimization of sub- merged culture conditions for mycelial growth and exo-biopolymer production by Auricularia polytricha by one-factor-at-a-time and uniform design (UD) meth- ods. First, the one-factor-at-a-time method was adop- ted to investigate the effects of environmental factors (i.e., initial pH and temperature) and variables of me- dium components (i.e., carbon, nitrogen and mineral sources) on mycelial growth and exo-biopolymer pro- duction. Sucrose, yeast extract, and K 2 HPO 4 were identified to be the most suitable carbon, nitrogen, and mineral sources, respectively. The optimal tem- perature and initial pH for mycelial growth and exo- biopolymer production were found to be 25 ◦ C and 5.0, respectively. Subsequently, the concentrations of sucrose, yeast extract, and K 2 HPO 4 were opti- mized using the UD method. The optimal concen- trations for the enhanced production were determined as 6 % (w/v) sucrose, 2.5 % (w/v) yeast extract, and 0.3 % (w/v) K 2 HPO 4 for mycelial yield, and 6 % (w/v) sucrose, 1.28 % (w/v) yeast extract, and 0.3 % (w/v) K 2 HPO 4 for exo-biopolymer production, respectively. Subsequent experiments confirmed the validity of the models. This optimization strategy in shake-flask culture led to a mycelial yield of 6.14 g/l, and exo- biopolymer production of 2.12 g/l, respectively, which were considerably higher than those obtained in the preliminary studies. By using the optimized medium, the maximum concentrations of mycelial biomass and exo-biopolymer in a 5 litre stirred-tank bioreactor indicated 35.3 g/l and 3.1 g/l, respectively. Introduction Auricularia polytricha (pepeiao or wood ears fungus) is one of the famous Chinese medicinal entomopathogenic fungi that invades and lives in the woodcut or fallen logs of several types of trees. A. polytricha resemble ears in shape and are gelatinous to leathery in texture and brownish-purple in colour. The protein, vitamin, and carbohydrate content of A. polytricha are reported to be higher than those of many vegetables and fruits. A. polytricha, therefore, it is an ideal food for dietetic prevention of hyperlipidaemia [1]. According to Chinese medicine, fruit bodies of entomo- pathogenic fungi are highly valued as medicinal herbs, owing to their various biological and pharmacological activities, such as immuno-stimulating and antitumour activities [2–4]. Kaneda and Tokuda [5] have already investigated the hypolipidaemic effect of the fruiting body of A. polytricha. Yang et al. [6] have reported the hypolipidaemic effect of exo-polymer from A. polytricha. Although many investigators have attempted to obtain the optimal submerged culture for exo-biopolymer production from several fungi [7,8], to the best of our knowledge, the nutritional requirements and environmental conditions for submerged culture of A. polytricha have not been demonstrated. Medium optimization by the one-factor-at-a-time method involves changing one independent variable (i.e., nutrients, temperature, pH, etc.) while fixing the others at certain levels. This method is not only time-consuming, but also often leads to an incomplete understanding of the behaviour of the system, resulting in confusion and a lack of predictive ability. Hence, as a more practical method, uniform design (UD) was employed to study relationships between medium components and their effect on mycelial growth and exo-biopolymer production. In the present study, the statistical experimental design was used to optimize the submerged culture conditions to simultaneously produce the mycelial biomass and exo- biopolymer by A. polytricha. In the first step, the one-factor- at-a-time method was used to investigate the effects of environmental factors (i.e., pH and temperature) and variables of medium composition (i.e., carbon, nitrogen Key words: Auricularia polytricha (pepeiao or wood ears fungus), exo-biopolymer, morphology, submerged culture. Abbreviations used: DPS, data processing system; PDA, potato dextrose agar; UD, uniform design; VVM, vol. · vol. −1 · min −1 . 1 To whom correspondence should be addressed (e-mail jwyun@daegu.ac.kr). C  2003 Portland Press Ltd