Processos microbiológicos e enzimáticos na síntese quimioenzimática de fragrâncias

Abstract

The use of biocatalytic sources in the synthesis of molecules with odoriferous properties was the main goal of this work. Five compounds with odoriferous properties were prepared: (i) (5)-1- phenylethanol; (ii) (R)-1-phenylethanol; (iii) (5)-1-phenylethyl acetate; (iv) (R)-1-phenylethyl acetate and (v) Mugetanol. Firstly, we have used the yeast Candida tropicalis CE017 as biocatalyst in the synthesis of (5)-1-phenylethanol fragrance. The yeast was used as free or immobilized cells and reaction conditions were optimi7ed in order to improve the yield and enantiomeric excess. The conditions as substrate quantity, pH, culture medium, temperature, inoculum quantity and resting cells were tested. The bioreduction of acetophenone using C. tropicalis produced (S)-1-phenylethanol as a maximum yield of 62% and enantiomeric excess of 97% in favor of S enantiomer. In addition, (S) and (R)-1-phenylethanol were obtained by bioreduction of acetophenone using commercial alcohol dehydrogenases (ADHs): ADH A, ADH CP, ADH LB, ADH PR2, ADH RS1 and ADH T. Bioreduction catalyzed by enzymes LB and PR2 produced the (R)-1-phenylethanol with conversions of 97% and 36%, respectively, and ee 2:99%. On the other hand, using enzymes T, CP, RS1 and A the (5)-1-phenylethanol was obtained with 6-88% of conversion and ee 299%. The preparation of (S) and (R)-1-phenylethyl acetates was performed by classic chemical process (acetylation or Mitsunobu reactions) from optically pure alcohols. (S)-1-phenylethyl acetate was obtained with 78% yield and 97% ee. Other fragrance with great interest in the aroma industry, as Mugetanol, was synthesized through three independent routes. In the route (a) the chemical reduction of 4- isopropylacetophenone produced the 1-(4-isopropylphenyl) ethanol with 90% yield. The kinetic resolution step was performed with vinyl acetate and lipases CAL-B or PSL-C I in THF. Both lipases showed stereopreference for the acetylation of the (R)-enantiomer, with E>200. In this case, the high resolution rate was obtained using PSL-C I (ees 2:99% with 96% yield; cep 98% with 88% yield; conversion of 50% and E>200, with 6 h of reaction). Then, the alcohol obtained by kinetic resolution catalyzed by PSL-C I was reacted with hydrogen with a catalytic amount of chloro-(1,5-hexadiene)rhodium, giving Mugetanol as a mixture of (S)-cis-trans with 88% yield and de of 64% in favour of cis-isomer. In the route (b) the alcohol dehydrogenases LB and PR2 produced the (R)-1-(4-isopropylphenyl) ethanol, with conversions of 76 and 17% and ee 2:99%, respectively, while (5)-1-(4-isopropylphenyl) ethanol, was produced in the presence of enzymes T, CP, RS1 and A with 65-96% of conversions and ee >99%. The chemical reduction to obtain the optically active 1-(4-isopropylphenyl) ethanol can lead to Mugetanol with 88% yield. In the route (c), the catalytic hydrogenation of 4-isopropylacetophenone produced a mixture with difficult separation: 4-(isopropylcyclohexyl)methylketone (16%), racemic Mugetanol (32%) and 1-(4-isopropylphenyl) ethanol (22%). Some racemic Mugetanol was recovered and the lipase- catalyzed kinetic resolution was studied with the enzymes CAL-B and PSL-C I. The Mugetanol was obtained with 91% yield as a mixture of cis-trans with de of 22% in favor of cis-isomer and enantiomeric excess 2:99% in favor of S enantiomer, when CAL-B was used. In the presence of PSL-C I, the Mugetanol was obtained with 93% yield as a mixture of cis-trans and de of 16% in favor of cis-isomer. For the cis-isomer ee was 60% in favor of S enantiomer and for trans-isomer the ee was 86% in favor of S enantiomer