SOYBEAN AND HEALTH - University of Macau Library

238Soybean and Health4. Inherent problems and solutions in strain improvementThe microbial metabolism of sterols has already been shown to be a promising means ofpreparing valuable steroids. During the second half of the twentieth century, manymicrobial strains were developed and tailored to synthesize many kinds of C19 and C22steroids using phytosterols as substrates (Szentirmai, 1990). So far, however, the productionof steroids from phytosterols by microbial transformation is still not widely used in thepharmaceutical industry. This may be because of two inherent problems in the phytosterolbiotransformation process: (1) The low water solubility of phytosterols can lead to poorbioavailability. (2) The final products are toxic to microbial cells. Many technologicalstrategies have been proposed to overcome these problems. These have been reviewed indetail by Malaviya and Fernandes (Fernandes, et al., 2003; Malaviya & Gomes, 2008). Here,we mainly focus on how to develop robust organisms that can withstand their ownproducts.4.1 Low water solubility and adaptive mechanismsMost steroids have solubility below 0.1 mM. Phytosterols are more hydrophobic, their watersolubility usually being no more than 1 μM. The poor solubility of phytosterols seriouslyretards their bioavailability because of inadequate mass transfer (Goetschel & Bar, 1992;Phase & Patil, 1994; Malaviya & Gomes, 2008). Many efforts have been carried out toimprove the dispersity and dissolubility of phytosterols in reaction media. Among these,surfactant-facilitated emulsification, favored for its practicality and effectiveness, is acommon means of enhancing the bioavailability of phytosterols (Fernandes, et al., 2003;Malaviya & Gomes, 2008). There are other strategies, but many of these seem inadequate inpractice. For example, organic solvent facilitated dissolution significantly raises thecomplexity of the process and production costs and also results in environmental pressures.It is also not very productive.The mechanism of sterol uptake by microbial cells has already been demonstrated: duringthe transformation process, microbial cells adhere to the surfaces of sterol particles, formingstable agglomerates. Sterol uptake takes place via the direct contact between the cells andthe sterol particles (Atrat, et al., 1991; Goetschel & Bar, 1992; Fernandes, et al., 2003). This isan adaptive mechanism that allows microorganisms to use minimally water-solublehydrocarbons as carbon sources. Like phytosterols, many polycyclic aromatic hydrocarbons(PAH), such as naphthalene and anthracene, also exhibit extremely low water solubility andare also poorly bioavailable (Johnsen & Karlson, 2004; Harms, et al., 2010a, 2010b). Manymicroorganisms have been shown to use these hydrophobic hydrocarbons as carbon sourcesthrough similar evolutionary adaptations (Miyata, et al., 2004; Heipieper, et al., 2010; Parales& Ditty, 2010; Harms, et al., 2010a). Generally, the microorganisms adapted to usehydrophobic hydrocarbons have the following physiological properties: (1) lipophilic cellwalls and adaptive changes in the surface properties allowing direct adhesion to thehydrophobic substrates, (2) high affinity uptake systems such as active transporters andmembrane-associated enzymes for initial degradation, (3) the ability to excretebiosurfactants or bioemulsifiers in order to increase the bioavailability of hydrophobichydrocarbons (Wick, et al., 2002; Heipieper, et al., 2010; Miyata, et al., 2004; Parales & Ditty,2010; Johnsen & Karlson, 2004; Perfumo, et al., 2010). Several studies regarding adaptiveresponses to PAH have been carried out with Gram-positive bacteria belonging to themycolate-containing bacteria families, including Mycobacterium, Rhodococcus,