SHORT COMMUNICATION Heterotrophic growth and lipid production of Chlorella protothecoides on glycerol John O’Grady • John A. Morgan Received: 8 July 2010 / Accepted: 6 October 2010 / Published online: 26 October 2010 Ó Springer-Verlag 2010 Abstract During the production of biodiesel, a signifi- cant amount of glycerol is generated which currently has little commercial value. A study on the growth and lipid production of Chlorella protothecoides using glycerol as the carbon source was performed to demonstrate the utility of recycling crude glycerol created during biodiesel pro- duction. Glycerol was examined as both the sole carbon source and in combination with glucose. Algae cultures grown on only glycerol in shake flasks showed a specific growth rate and final lipid yield of 0.1/h and 0.31 g lipid/g substrate, respectively. The values were similar to those observed on pure glucose, 0.096/h and 0.24 g lipid/g sub- strate. When the media contained a mixture of glycerol and glucose, simultaneous uptake of the two substrates was observed. Due to the difference in rates of lipid storage, lipid production was 0.077 g lipid/(l h) during growth on glycerol, while growth on glucose had a productivity of 0.096 g lipid/(l h). During growth on the 9:1 mixture of both glucose and glycerol, lipid productivity was 0.098 g lipid/(l h). In order to simulate the use of waste glycerol from biodiesel production the experiments were repeated and similar growth rates, yields, and lipid productivities were achieved. Therefore, we have demonstrated the promise for simultaneous high growth rates and lipid yields of C. protothecoides heterotrophically grown on mixtures of glycerol. Keywords Glycerine  Glycerin  Algae Introduction In view of increasing energy costs and the global impact of petroleum based fuel, focus on developing alternative energy solutions from renewable resources is growing. For liquid fuel, biodiesel is promising because it can be utilized by current engines without requiring modifications to the design or materials [1]. Microalgae are considered one of the most promising sources of biodiesel [2]. Microalgae, unlike most sources of biofuels, may be grown on non- arable land, or in fresh or salt water, avoiding competition with traditional food and feed crops [3]. Multiple strains of algae have been shown to accumulate lipids to as much as 80% of their dry weight [3, 4] when grown heterotrophi- cally in nitrogen deficient conditions. Heterotrophic con- ditions can also significantly increase growth rates and final cell densities compared to autotrophic conditions [5]. These factors alone, however, do not offset the higher costs of heterotrophic algae production. Foremost among these costs is the cost of the substrate on which the algae are grown. Multiple studies have focused on ways to make bio- diesel from microalgae more economical. One of the methods examined is to decrease substrate costs by using alternative carbon sources: corn powder hydrolysate [6], sweet sorghum juice [7], or wastewater with high bio- availability [2, 8]. Alternatively, studies have sought to find marketable uses for the byproducts of algal biodiesel production, such as glycerol [9]. Others have tried to produce specialty chemicals in algae while simultaneously growing them for biodiesel production [2]. A primary issue for biodiesel production is to maintain both lipid yield and growth rates at or near the levels observed on glucose while decreasing the costs by employing a dif- ferent substrate. J. O’Grady  J. A. Morgan (&) School of Chemical Engineering, Purdue University, FRNY Hall, West Lafayette, IN 47907-2050, USA e-mail: jamorgan@purdue.edu 123 Bioprocess Biosyst Eng (2011) 34:121–125 DOI 10.1007/s00449-010-0474-y