Production of 2‐phenylethanol and 2‐phenylethylacetate from L‐phenylalanine by coupling whole‐cell biocatalysis with organophilic pervaporation

Abstract

An integrated bioprocess for the production of the natural rose-like aroma compounds, 2-phenylethanol (2-PE) and 2-phenylethylacetate (2-PEAc), from L-phenylalanine (L-phe) with yeasts was investigated. The hydrophobicity of the products leads to product inhibition, which can be compensated by in situ product removal (ISPR). An organophilic pervaporation unit, equipped with a polyoctylmethylsiloxane (POMS) membrane, was coupled via a bypass to a bioreactor and proved to be a suitable technique for the in situ removal of high-boiling products from culture broth. With batch cultures of the thermotolerant yeast Kluyveromyces marxianus CBS 600 in a standard medium at 35 degrees C, the use of pervaporation resulted in a double 2-PE concentration (2.2 g/L) and 1.3 g/L 2-PEAc, which only accumulated transiently in low concentrations during cultivation without ISPR. Using a previously optimized medium, the variation of the temperature from 30 degrees C to 40 degrees C caused an increase in the total conversion yield from 63% to 79%, corresponding to total product concentrations of 5.23 and 5.85 g/L, respectively. In the 40 degrees C batch experiment, the volumetric productivity (2-PE + 2-PEAc) during the exponential phase was 5.2 mmol/L h. While for 2-PE, there is still potential for further optimization, the more hydrophobic 2-PEAc was nearly completely removed from the aqueous culture broth (enrichment factor >400), resulting in highly aroma-enriched permeates. Due to the temperature-correlated performance of the pervaporation, the bioconversion was still efficient even at 45 degrees C (conversion yield: 69%). Surprisingly, at 45 degrees C, the molar ratio of the two products inverted and 2-PEAc turned out to be the main product (4.0 g/L), which opens easy control of the reaction's selectivity by external means. Retrofitting the process with interim heating and cooling equipment to use different temperature levels for cultivation and pervaporation resulted in a decreased yield and product concentration caused by multiple stress factors. The medium composition affected the pervaporation efficiency with molasses acting detrimental.

Comparing in situ removal strategies for improving styrene bioproduction

“…Examples of successful approaches for the in situ recovery of inhibitory biomolecules include solvent extraction [20,23,24], adsorption [25][26][27], gas stripping [28], vacuum stripping [29], and membrane pervaporation [30,31]. Meanwhile, whereas in situ recovery strategies have been demonstrated as effective for improving the bioproduction of other aromatic compounds (e.g., 2-phenylethanol, phydroxystyrene, (S)-styrene oxide, and benzaldehyde) [32][33][34][35][36][37], their application to bio-derived styrene has not yet been reported. Moreover, whereas said prior studies have predominantly focused on single step biotransformations, the alternative focus here on de novo biosynthesis directly from renewable glucose presents new challenges.…”

Lipase-catalysed synthesis of natural aroma-active 2-phenylethyl esters in coconut cream

“…The production of 2-phenylethanol and its ester 2-phenylethyl acetate by various yeasts has been relatively well studied (Alber- Cardillo, Servi, & Zucchi, 1994;Etschmann, Bluemke, Sell, & Schrader, 2002;Etschmann & Schrader, 2006;Etschmann, Sell, & Schrader, 2003, 2005Huang, Lee, & Chou, 2000;Stark, Munch, Sonnleitner, Marison, & Stockar von, 2002;Wittmann, Hans, & Bluemke, 2002). However, the production of other aroma-active, yet commercially valuable esters of 2-phenylethanol, such as 2-phenylethyl esters of C3-C8 fatty acids, has received no or little research attention.…”

Bioproduction of the aroma compound 2‐Phenylethanol in a solid–liquid two‐phase partitioning bioreactor system by Kluyveromyces marxianus

“…Etschmann and Schrader (2006) achieved the highest 2-PE concentration (10.2 g/L overall) reported by using K. marxianus CBS 600. Other ISPR techniques such as microcapsulation (Serp et al, 2003;Stark et al, 2003); and organophilic pervaporation (Etschmann et al, 2005) have also been reported.…”