6
A.B. Martins et al. / Ultrasonics Sonochemistry xxx (2013) xxx–xxx
Novozym 435 for flavor ester synthesis in three ways: (a) higher
biocatalyst operational stability, (b) biocatalyst stabilization in
the presence of high concentrations of acetic acid in n-hexane,
and (c) increased productivity of ester, when compared to the tra-
ditional method of mechanical agitation. The reaction parameters
were optimized and the optimal conditions were similar to
mechanical agitation. In 2 h was possible to obtain around 95% of
yield conversion at 0.3 M acetic acid. Nevertheless, in spite of the
similarity in the reaction conditions, the ultrasound-assisted tech-
nology allowed the direct reuse of the enzyme without the needed
n-hexane washings. The enzyme could be reused, keeping around
[5] S. Serra, C. Fuganti, E. Brenna, Biocatalytic preparation of natural flavours and
fragrances, Trends Biotechnol. 23 (2005) 193–198.
[
6] H. Abbas, L. Comeau, Aroma synthesis by immobilized lipase from Mucor sp,
Enzyme Microb. Technol. 32 (2003) 589–595.
[7] R. Ben Salah, H. Ghamghui, N. Miled, H. Mejdoub, Y. Gargouri, Production of
butyl acetate ester by lipase from novel strain of Rhizopus oryzae, J. Biosci.
Bioeng. 103 (2007) 368–372.
[
8] M. Karra-Chaabouni, H. Ghamgui, S. Bezzine, A. Rekik, Y. Gargouri, Production
of flavour esters by immobilized Staphylococcus simulans lipase in a solvent-
free system, Process Biochem. 41 (2006) 1692–1698.
[
9] G.V. Kumar, M.N. Rao, Enzymatic synthesis of butyl butyrate using response
surface methodology, J. Food Sci. Technol. 41 (2004) 560–562.
[10] G.D. Yadav, P.S. Lathi, Kinetics and mechanism of synthesis of butyl
isobutyrate over immobilised lipases, Biochem. Eng. J. 16 (2003) 245–252.
11] E.V. Rokhina, P. Lens, J. Virkutyte, Low-frequency ultrasound in biotechnology:
state of the art, Trends Biotechnol. 27 (2009) 298–306.
[
7
0% of its original activity after 14 reaction cycles, without any
treatment. This represents a reduction in the generated residues,
and save one unit operation in the process, decreasing the cost of
production. Under ultrasound irradiation, it was possible to use
up to 2.5 M acetic acid, increasing the process productivity. There-
fore, improving the acid concentration, the number of batches
reuses, avoiding the unit operations of washing to recover the en-
zyme activity, and the fact that ultrasound requires only one-third
to half the energy consumed by mechanical agitation, it can be sta-
ted that ultrasound-assisted butyl acetate synthesis catalyzed by
Novozym 435 could be scaled up to industrial size in a near future.
Further research aiming at determining whether or not varying
ultrasound frequency will have any influence on the reaction rate
are granted.
[12] C.B. Hobuss, D. Venzke, B.S. Pacheco, A.O. Souza, M.A.Z. Santos, S. Moura, F.H.
Quina, K.G. Fiametti, J. Vladimir Oliveira, C.M.P. Pereira, Ultrasound-assisted
synthesis of aliphatic acid esters at room temperature, Ultrason. Sonochem. 19
(
2012) 387–389.
[13] B. Kwiatkowska, J. Bennett, J. Akunna, G.M. Walker, D.H. Bremner, Stimulation
of bioprocesses by ultrasound, Biotechnol. Adv. 29 (2011) 768–780.
[
14] M.M. Zheng, L. Wang, F.H. Huang, L. Dong, P.M. Guo, Q.C. Deng, W.L. Li, C.
Zheng, Ultrasonic pretreatment for lipase-catalyed synthesis of phytosterol
esters with different acyl donors, Ultrason. Sonochem. 19 (2012) 1015–1020.
[15] J.V. Sinisterra, Application of ultrasound to biotechnology: an overview,
Ultrasonics 30 (1992) 180–185.
[
16] L. Batistella, L.A. Lerin, P. Brugnerotto, A.J. Danielli, C.M. Trentin, A. Popiolski, H.
Treichel, J.V. Oliveira, D. De Oliveira, Ultrasound-assisted lipase-catalyzed
transesterification of soybean oil in organic solvent system, Ultrason.
Sonochem. 19 (2012) 452–458.
[
[
[
17] V.G. Deshmane, P.R. Gogate, A.B. Pandit, Process intensification of synthesis
process for medium chain glycerides using cavitation, Chem. Eng. J. 145 (2008)
351–354.
18] V.G. Deshmane, P.R. Gogate, A.B. Pandit, Ultrasound assisted synthesis of
isopropyl esters from palm fatty acid distillate, Ultrason. Sonochem. 16 (2009)
Acknowledgments
345–350.
19] A.B. Martins, N.G. Graebin, A.S.G. Lorenzoni, R. Fernandez-Lafuente, M.A.Z.
Ayub, R.C. Rodrigues, Rapid and high yields of synthesis of butyl acetate
catalyzed by Novozym 435: reaction optimization by response surface
methodology, Process Biochem. 46 (2011) 2311–2316.
This work was supported by grants from Fundação de Amparo a
Pesquisa do Rio Grande do Sul (FAPERGS – ARD/2011), from CNPq
(
Brazilian Bureau of Science and Technology), and CTQ2009-07568
[
[
20] N.G. Graebin, A.B. Martins, A.S.G. Lorenzoni, C. Garcia-Galan, R. Fernandez-
Lafuente, M.A.Z. Ayub, R.C. Rodrigues, Immobilization of lipase B from Candida
antarctica on porous styrene-divinylbenzene beads improves butyl acetate
synthesis, Biotechnol. Prog. 28 (2012) 406–412.
21] M.D. Romero, L. Calvo, C. Alba, A. Daneshfar, H.S. Ghaziaskar, Enzymatic
synthesis of isoamyl acetate with immobilized Candida antarctica lipase in n-
hexane, Enzyme Microb. Technol. 37 (2005) 42–48.
from Spanish Ministerio de Ciencia e Innovación. The authors
would like to thank Mr. Ramiro Martínez (Novozymes, Spain) for
kindly supplying the enzymes used in this research. We also thank
CNPq – Brazil for a fellowship to J.L.R. Friedrich and FAPERGS –
Brazil for a fellowship to A.B. Martins.
[
[
22] D. Wei, C. Gu, Q. Song, W. Su, Enzymatic esterification for glycoside lactate
synthesis in organic solvent, Enzyme Microb. Technol. 33 (2003) 508–512.
23] J.L.R. Friedrich, F.P. Peña, C. Garcia-Galan, R. Fernandez-Lafuente, M.A.Z. Ayub,
R.C. Rodrigues, Effect of immobilization protocol on optimal conditions of
References
[
[
[
1] A. Rajendran, A. Palanisamy, V. Thangavelu, Lipase catalyzed ester synthesis for
food processing industries, Braz. Arch. Biol. Technol. 52 (2009) 207–219.
2] K.-E. Jaeger, T. Eggert, Lipases for biotechnology, Curr. Opin. Biotechnol. 13
[24] C.-H. Kuo, S.-H. Chiang, H.-Y. Ju, Y.-M. Chen, M.-Y. Liao, Y.-C. Liu, C.-J. Shieh,
Enzymatic synthesis of rose aromatic ester (2-phenylethyl acetate) by lipase, J.
Sci. Food Agric. 92 (2012) 2141–2147.
[25] V.B. Veljkovi c´ , J.M. Avramovi c´ , O.S. Stamenkovi c´ , Biodiesel production by
ultrasound-assisted transesterification: State of the art and the perspectives,
Renew. Sustain. Energy Rev. 16 (2012) 1193–1209.
(
2002) 390–397.
3] S. Torres, M.D. Baigori, S.L. Swathy, A. Pandey, G.R. Castro, Enzymatic synthesis
of banana flavour (isoamyl acetate) by Bacillus licheniformis S-86 esterase, Food
Res. Int. 42 (2009) 454–460.
[
4] A.R.M. Yahya, W.A. Anderson, M. Moo-Young, Ester synthesis in
lipase-catalyzed reactions, Enzyme Microb. Technol. 23 (1998) 438–450.