dbbaf0aa7db8c08b167affbbb1045f37 catalysts_a2022v12n6p639.pdf 29c2030535b7de933851629f36b46f7cc672d235 catalysts_a2022v12n6p639.pdf d8377e0c76c133abd9b4541653ded418bd9292e97a38c3c367fb1da968f07124 catalysts_a2022v12n6p639.pdf Title: Producing Natural Flavours from Isoamyl Alcohol and Fusel Oil by Using Immobilised Rhizopus oryzae Lipase Subject: Enzymatic synthesis of short-chain esters (flavours) might enable their labelling as natural, increasing their value. Covalently immobilised Rhizopus oryzae lipase (EO-proROL) was used to synthesise isoamyl butyrate and acetate. In cyclohexane, the best performer reaction solvent, 1.8 times higher yield of isoamyl butyrate (ca. 100%) than isoamyl acetate (ca. 55%) was obtained. Optimum initial acid concentration (410 mM) and acid:alcohol mole ratio (0.5) were established by a central composite rotatable design to maximise isoamyl butyrate single-batch and cumulative production with reused enzyme. These conditions were used to scale up the esterification (150 mL) and to assess yield, initial esterification rate, productivity and enzyme operational stability. Commercial isoamyl alcohol and fusel oil results were found to be similar as regards yield (91% vs. 84%), initial reaction rate (5.4 M min-1 with both substrates), operational stability (40% activity loss after five runs with both) and productivity (31.09 vs. 28.7 mM h-1). EO-proROL specificity for the structural isomers of isoamyl alcohol was also evaluated. Thus, a successful biocatalyst and product conditions ready to be used for isoamyl ester industrial production are here proposed. Keywords: Rhizopus oryzae lipase; flavor; fusel oil; short-chain esters; esterification; Komagataella phaffii; immobilization; isoamyl alcohol; specificity Author: Josu López-Fernández, Maria Dolors Benaiges, Xavier Sebastian, Jose María Bueno and Francisco Valero Creator: LaTeX with hyperref Producer: pdfTeX-1.40.21 CreationDate: Wed Jun 15 08:49:27 2022 CEST ModDate: Wed Jun 15 08:53:19 2022 CEST Tagged: no UserProperties: no Suspects: no Form: none JavaScript: no Pages: 13 Encrypted: no Page size: 595.276 x 841.89 pts (A4) Page rot: 0 File size: 1818089 bytes Optimized: no PDF version: 1.7 name type encoding emb sub uni object ID ------------------------------------ ----------------- ---------------- --- --- --- --------- AICEPB+VnURWPalladioL Type 1 Custom yes yes no 10 0 LEVXPH+EURM10 Type 1 Builtin yes yes no 15 0 DLNHUS+URWPalladioL-Bold Type 1 Custom yes yes no 19 0 AYBNBX+URWPalladioL-Roma Type 1 Custom yes yes no 24 0 AORCJL+URWPalladioL-Ital Type 1 Custom yes yes no 28 0 UPACYV+URWPalladioL-BoldItal Type 1 Custom yes yes no 32 0 VORBJF+CMSY10 Type 1 Builtin yes yes no 36 0 PRJKGQ+VnURWPalladioL-Bold Type 1 Custom yes yes no 40 0 CGDLTM+TimesNewRomanPS-BoldItalicMT Type 1C WinAnsi yes yes no 58 0 XIRVNA+EURB10 Type 1 Builtin yes yes no 89 0 Jhove (Rel. 1.22.1, 2019-04-17) Date: 2023-05-06 02:26:47 CEST RepresentationInformation: catalysts_a2022v12n6p639.pdf ReportingModule: PDF-hul, Rel. 1.12.1 (2019-04-17) LastModified: 2022-06-23 11:39:40 CEST Size: 1818089 Format: PDF Version: 1.7 Status: Well-Formed and valid SignatureMatches: PDF-hul MIMEtype: application/pdf PDFMetadata: Objects: 361 FreeObjects: 1 IncrementalUpdates: 0 DocumentCatalog: PageLayout: SinglePage PageMode: UseNone Outlines: Item: Title: Introduction Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@44c8afef Item: Title: Results and Discussion Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@7b69c6ba Children: Item: Title: Reaction Solvent for Butyric and Acetic Acid Esterification with Isoamyl Alcohol Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@46daef40 Item: Title: Experimental Design: Optimisation of Isoamyl Butyrate Single-Batch and Cumulative Production Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@12f41634 Item: Title: Reaction Scale Up: Isoamyl Butyrate Production, Fusel Oil Use and Half-Lives Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@13c27452 Item: Title: Structural Isomers: 3-Methylbutanol and 2-Methylbutanol Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@262b2c86 Item: Title: Materials and Methods Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@371a67ec Children: Item: Title: Chemicals Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@5ed828d Item: Title: Lipase Production and Downstream Processing Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@50d0686 Item: Title: Lipase Activity Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@7a3d45bd Item: Title: Lipase Immobilisation Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@1e7c7811 Item: Title: Esterification Reactions Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@77ec78b9 Children: Item: Title: Influence of the Acid Concentration and Acid:Alcohol Mole Ratio Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@1a3869f4 Item: Title: Reaction Scale Up Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@a38d7a3 Item: Title: Statistical Analysis Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@77f99a05 Item: Title: Gas Chromatography/Mass Spectrometry Analysis Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@63440df3 Item: Title: Conclusions Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@3aeaafa6 Item: Title: References Destination: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject@76a3e297 Info: Title: Producing Natural Flavours from Isoamyl Alcohol and Fusel Oil by Using Immobilised Rhizopus oryzae Lipase Author: Josu López-Fernández, Maria Dolors Benaiges, Xavier Sebastian, Jose María Bueno and Francisco Valero Subject: Enzymatic synthesis of short-chain esters (flavours) might enable their labelling as natural, increasing their value. Covalently immobilised Rhizopus oryzae lipase (EO-proROL) was used to synthesise isoamyl butyrate and acetate. In cyclohexane, the best performer reaction solvent, 1.8 times higher yield of isoamyl butyrate (ca. 100%) than isoamyl acetate (ca. 55%) was obtained. Optimum initial acid concentration (410 mM) and acid:alcohol mole ratio (0.5) were established by a central composite rotatable design to maximise isoamyl butyrate single-batch and cumulative production with reused enzyme. These conditions were used to scale up the esterification (150 mL) and to assess yield, initial esterification rate, productivity and enzyme operational stability. Commercial isoamyl alcohol and fusel oil results were found to be similar as regards yield (91% vs. 84%), initial reaction rate (5.4 M min-1 with both substrates), operational stability (40% activity loss after five runs with both) and productivity (31.09 vs. 28.7 mM h-1). EO-proROL specificity for the structural isomers of isoamyl alcohol was also evaluated. Thus, a successful biocatalyst and product conditions ready to be used for isoamyl ester industrial production are here proposed. 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