Rhodotorula minuta

The process uses cells of Rhodotorula minuta entrapped in polyurethane. These cells selectively hydrolyse the L-ester. The remaining D-menthyl succinate is then hydrolysed and the 15-menthol racemised via D-menthone and then recycled. We have represented this process in Figure 9.9. 

Figure 9.9 Resolution of DL-menthol using an esterase from Rhodotorula minuta var texensis.

The L-menthol could be recovered by hydrolysing the L-menthyl-5-phenylvalerate. You might anticipate using an esterase to carry out this hydrolysis such as the enzyme from Rhodotorula minuta described in the text. 

A cytochrome P450 has been purified from Saccharomyces cerevisiae that has benzo[a]pyrene hydroxylase activity (King et al. 1984), and metabolizes benzo[fl]pyrene to 3- and 9-hydroxybenzo[fl]pyrene and benzo[fl]pyrene-7,8-dihydrodiol (Wiseman and Woods 1979). The transformation of PAHs by Candida Upolytica produced predominantly monohydroxyl-ated products naphth-l-ol from naphthalene, 4-hydroxybiphenyl from biphenyl and 3- and 9-hydroxybenzo[fl]pyrene from benzo[fl]pyrene (Cerniglia and Crow 1981). The transformation of phenanthrene was demonstrated in a number of yeasts isolated from littoral sediments and of these, Trichosporumpenicillatum was the most active. In contrast, biotransformation of benz[fl]anthracene by Candida krusei and Rhodotorula minuta was much slower (MacGillivray and Shiaris 1993). 

Detailed studies have been made on the /3-D-glucosidases of the following (t) Rhodotorula minuta (misnamed Saccharomyces cerevi-stae),32-335,338 (it) Kluyveromyces (Saccharomyces) lactis,337-344 (tit) Kluyveromyces (fragilis) marxianus,348,348 (iv) Kluyveromyces dobz-hanskii,348 (o) a hybrid of Kluyveromyces marxianus and Kluyveromyces dobzhanskii,348-351 (t>t) a yeast of unknown identity (see Ref. 4) called Saccharomyces cerevisiae, 36 3M-3 8 (mi) Pichia (Candida) guilliermondii,331 and (viii) Debaryomyces polymorphus (Pichia poly-morpha).358 Table X lists some characteristics of these /3-D-glucosi-... 

Rh, Rhodotorula minuta, Refs. 32 and 335 Kl, Kluyveromyces interspecific hybrid, Ref. 347 U, unidentified yeast (Refs. 355 and 356) D(i), D(ii), two /3-D-glucosidases of a strain of Debaryomyces polymorphus (Ref. 358). Induced enzymic activity, uninduced enzymic activity.c The relative rates are calculated from values of V. 

Debaryomyces polymorphus strain CBS 4349 Rhodotorula minuta strain CBS 2179 Candida tsukubaensis strain CBS 6389... 

Some detailed studies have been made on /3-D-glucosidases of the following (i) Rhodotorula minuta (misnamed Saccharomyces cerevisiae ),538,539 (it) Kluyveromyces (Saccharomyces) lactis,540-546 (f )... 

Rhodotorula mucilaginosa, Rh. minuta, Sporobolomyces albidus, and Sp. salmonicolor, but present in Cryptococcus diffluens. However, Meyer (49) clearly demonstrated such activity in the intracellular extract and in the cell walls of Rhodotorula minuta var. texensis, Rh. glutinis, Cryptococcus albidus, Cr. terreus, and Cr. laurentii. The activity in the cell walls of these yeasts was always much higher than that of the intracellular extract. 0-(l- 6)-Glucanase activity of these species was either lacking or extremely low, but chitinase activity was significant in most strains. 

Fukuda, H., T. Fujii, E. Sukita, M. Tazaki, S. Nagahama, and T. Ogawa (1994). Reconstitution of the isobutene-forming reaction catalyzed by cytochrome P450 and P450 reductase from Rhodotorula minuta Decarboxylation with the formation of isobutene. Biochem. Biophys. Res. Commun. 201, 516-522. 

Fukuda, H., K. Nakamura, E. Sukita, T. Ogawa, and T. Fujii (1996). Cytochrome P450rm from Rhodotorula minuta catalyzes 4-hydroxylation of benzoate. J Biochem. 119, 314-318. 

Omata, T., N. Iwamoto, T. Kimura, A. Tanaka, S. Fukui, 1981. Stereoselective hydrolysis of <7/-menthyl succinate by gei-entrapped Rhodotorula minuta var. texensis cells in organic solvent. Appl. Microbiol. BiotechnoL, 11 119-204. 

In this example, synthetic DJ.-menthol racemate is initially chemically esterified with acetate, formate, propionate, myristate, benzoate, or succinate. The ester is then selectively hydrolyzed by steriospecilic microbial enzymes to produce L-menthol that is readily isolated from the D-menthol ester. The low water solubility of the menthyl esters and L-menthol has made this process particularly suitable to non-aqueous systems. Omata et al. [90] immobilized cells of Rhodotorula minuta in photo-cross-linked or polyurethane resin gels and used a water saturated hexane eluant to obtain a product of 100% optical purity. The immobilized cells had an estimated half-life of 55 to 63 days in this solvent. 

Omata, T., Stereoselective hydrolysis of D,L menthyl succinate by gel-entrapped Rhodotorula minuta var. texensis in organic solvent, Ap/>/. Microbiol. Biotechnol, 11, p. 199, 1981. 

Fujii et al. (1988) reported a mechanism of isobutene formation by the yeast Rhodotorula minuta in a culture medium that contained branched-chain carboxylic or amino acids. It was suggested that the pathway leads over isovalerate that is decarboxylated to isobutene. 

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