1.Enzymes in Fruit and Vegetable Processing Chemistry and Engineering Applications 94335.indb 1 3/31/10 4:29:03 PM 2010 Taylor and Francis Group, LLC

2. 94335.indb 2 3/31/10 4:29:04 PM 2010 Taylor and Francis Group, LLC 3. CRC Press is an imprint of the Taylor & Francis Group, an informa business Boca Raton London New York Enzymes in Fruit and Vegetable Processing Chemistry and Engineering Applications 94335.indb 3 3/31/10 4:29:04 PM 2010 Taylor and Francis Group, LLC 4. CRC Press Taylor & Francis Group 6000 Broken Sound Parkway NW, Suite 300 Boca Raton, FL 33487-2742 2010 by Taylor and Francis Group, LLC CRC Press is an imprint of Taylor & Francis Group, an Informa business No claim to original U.S. Government works Printed in the United States of America on acid-free paper 10 9 8 7 6 5 4 3 2 1 International Standard Book Number-13: 978-1-4200-9434-3 (Ebook-PDF) This book contains information obtained from authentic and highly regarded sources. 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Visit the Taylor & Francis Web site at http://www.taylorandfrancis.com and the CRC Press Web site at http://www.crcpress.com 5. v Contents Preface................................................................................................................ vii The Editor............................................................................................................ix List of Contributors............................................................................................xi 1Chapter Introduction to Enzymes............................................................1 Alev Bayndrl 2Chapter Effect of Enzymatic Reactions on Color of Fruits and Vegetables............................................................................19 J. Brian Adams 3Chapter Major Enzymes of Flavor Volatiles Production and Regulation in Fresh Fruits and Vegetables..........................45 Jun Song 4Chapter Effect of Enzymatic Reactions on Texture of Fruits and Vegetables............................................................................71 Luis F. Goulao, Domingos P. F. Almeida, and Cristina M. Oliveira 5Chapter Selection of the Indicator Enzyme for Blanching ofVegetables.............................................................................123 Vural Gkmen 6Chapter Enzymatic Peeling of Citrus Fruits...................................... 145 Maria Teresa Pretel, Paloma Snchez-Bel, Isabel Egea, andFelix Romojaro 7Chapter Use of Enzymes for Non-Citrus Fruit Juice Production................................................................................. 175 Liliana N. Ceci and Jorge E. Lozano 94335.indb 5 3/31/10 4:29:06 PM 2010 Taylor and Francis Group, LLC 6. vi Contents 8Chapter Enzymes in Citrus Juice Processing....................................197 Domenico Cautela, Domenico Castaldo, Luigi Servillo, andAlfonso Giovane 9Chapter Use of Enzymes for Wine Production..................................215 Encarna Gmez-Plaza, Inmaculada Romero-Cascales, andAna Beln Bautista-Ortn 1Chapter 0 Effect of Novel Food Processing on Fruit and Vegetable Enzymes..........................................................245 Indrawati Oey 1Chapter 1 Biosensors for Fruit and Vegetable Processing.................313 Danielle Cristhina Melo Ferreira, Lucilene Dornelles Mello, Renata Kelly Mendes, and Lauro Tatsuo Kubota 1Chapter 2 Enzymes in Fruit and Vegetable Processing: Future Trends in Enzyme Discovery, Design, Production, and Application........................................................................341 Marco A. van den Berg, Johannes A. Roubos, and Lucie Parenicov Index.................................................................................................................359 94335.indb 6 3/31/10 4:29:06 PM 2010 Taylor and Francis Group, LLC 7. vii Preface Fruits and vegetables are consumed as fresh or processed into different type of products. Some of the naturally occurring enzymes in fruits and vegetables have undesirable effects on product quality, and therefore enzyme inactivation is required during fruit and vegetable processing in order to increase the product shelf-life. Commercial enzyme prepara- tions are also used as processing aids in fruit and vegetable processing to improve the process efficiency and product quality, because enzymes show activity on specific substrates under mild processing conditions. Therefore, there has been a striking growth in the enzyme market for the fruit and vegetable industry. While fruit and vegetable processing is the subject of many books and other publications, the purpose of this book is to give detailed infor- mation about enzymes in fruit and vegetable processing from chemistry to engineering applications. Chapters are well written by an authorita- tive author(s) and follow a consistent style. There are 12 chapters in this book, and the chapters provide a comprehensive review of the chapter title important to the field of enzymes and fruit and vegetable processing by focusing on the most promising new international research develop- ments and their current and potential industry applications. Fundamental aspects of enzymes are given in Chapter 1. Color, flavor, and texture are important post-harvest quality parameters of fruits and vegetables. There are a number of product-specific details, dependent on the morphology, composition, and character of the individual produce. Chapters 2, 3, and 4 describe in detail the effect of enzymes on color, flavor, and texture of selected fruits and vegetables. Selection of the indicator enzyme for blanch- ing of vegetables is summarized in Chapter 5. For enzymes as processing aids, Chapter 6 describes in detail the enzymatic peeling of citrus fruits and Chapter 7 presents the importance of enzymes for juice production from pome, stone, and berry fruits. Inactivation of enzymes is required to obtain cloudy juice from citrus fruits. Chapter 8 is related to citrus juices; orange juice receives particular attention. Enzymes also play an impor- tant role in winemaking. The application of industrial enzyme prepara- tions in the wine industry is a common practice. The use of enzymes for 94335.indb 7 3/31/10 4:29:06 PM 2010 Taylor and Francis Group, LLC 8. viii Preface wine production is the focus of Chapter 9. Chapter 10 provides serious review of the inactivation effect of novel technologies on fruit and veg- etable enzymes to maximize product quality. Chapter 11 presents both chemical and technological information on enzyme-based biosensors for fruit and vegetable processing. The literature reported in each chap- ter highlights the current status of knowledge in the related area. Future trends for industrial use of enzymes are discussed in Chapter 12. The conclusion part of each chapter also presents the reader with potential research possibilities and applications. This book is a reference book to search or learn more about fruit and vegetable enzymes and enzyme-based processing of fruit and veg- etables according to the latest enzyme-assisted technologies and poten- tial applications of new approaches obtained from university and other research centers and laboratories. Such knowledge is important for the companies dealing with fruit and vegetable processing to be com- petitive and also for the collaboration among industry, university, and research centers. This book is also for the graduate students and young researchers who will play an important role for future perspectives of enzymes in fruit and vegetable processing. Alev Bayndrl 94335.indb 8 3/31/10 4:29:07 PM 2010 Taylor and Francis Group, LLC 9. ix The Editor Alev Bayndrl is a professor in the Department of Food Engineering, Middle East Technical University, Ankara, Turkey. She has authored or co-authored 30 journal articles. She received a BS degree (1982) from the Department of Chemical Engineering, Middle East Technical University. MS (1985) and PhD (1989) degrees are from the Department of Food Engineering, Middle East Technical University. She is work- ing on food chemistry and technology, especially enzymes in fruit and vegetable processing. 94335.indb 9 3/31/10 4:29:07 PM 2010 Taylor and Francis Group, LLC 10. 94335.indb 10 3/31/10 4:29:07 PM 2010 Taylor and Francis Group, LLC 11. xi List of Contributors J. Brian Adams Formerly of Campden & Chorleywood Food Research Association (Campden BRI) Chipping Campden Gloucestershire, UK Domingos P.F. Almeida Faculdade de Cincias Universidade do Porto Porto, Portugal Escola Superior de Biotecnologia Universidade Catlica Portuguesa Porto, Portugal Ana Beln Bautista-Ortn Departamento de Tecnologa de Alimentos, Nutricin y Bromatologa Facultad de Veterinaria, Universidad de Murcia Campus de Espinardo Murcia, Spain Alev Bayndrl Middle East Technical University Department of Food Engineering Ankara, Turkey Inmaculada Romero-Cascales Departamento de Tecnologa de Alimentos, Nutricin y Bromatologa Facultad de Veterinaria, Universidad de Murcia Campus de Espinardo Murcia, Spain Domenico Castaldo Stazione Sperimentale per le Industrie delle Essenze e dei Derivati dagli Agrumi (SSEA) Reggio Calabria, Italy Domenico Cautela Stazione Sperimentale per le Industrie delle Essenze e dei Derivati dagli Agrumi (SSEA) Reggio Calabria, Italy Liliana N. Ceci Planta Piloto de Ingeniera Qumica UNS-CONICET Baha Blanca, Argentina Isabel Egea Departamento Biologa del Estrs y Patologa Vegetal Centro de Edafologa y Biologa Aplicada del Segura-CSIC Espinardo, Murcia, Spain 94335.indb 11 3/31/10 4:29:07 PM 2010 Taylor and Francis Group, LLC 12. xii List of Contributors Danielle Cristhina Melo Ferreira Institute of Chemistry Unicamp Campinas, So Paulo, Brazil Alfonso Giovane Dipartimento di Biochimica e Biofisica Seconda Universit degli Studi di Napoli Napoli, Italy Vural Gkmen Department of Food Engineering Hacettepe University Ankara, Turkey Encarna Gmez-Plaza Departamento de Tecnologa de Alimentos, Nutricin y Bromatologa Facultad de Veterinaria, Universidad de Murcia Campus de Espinardo Murcia, Spain Luis F. Goulao Seco de Horticultura Instituto Superior de Agronomia Lisbon, Portugal Centro de Ecofisiologia, Bioquimica e Biotecnologia Vegetal Instituto de Investigao Cientifica Tropical Oeiras, Portugal Lauro Tatsuo Kubota Institute of Chemistry Unicamp Campinas, So Paulo, Brazil Jorge E. Lozano Planta Piloto de Ingeniera Qumica UNS-CONICET Baha Blanca, Argentina Lucilene Dornelles Mello UNIPAMPA Campus Bag Bag, RS, Brazil Renata Kelly Mendes Institute of Chemistry Unicamp Campinas, So Paulo, Brazil Indrawati Oey Department of Food Science University of Otago Dunedin, New Zealand Cristina M. Oliveira Seco de Horticultura Instituto Superior de Agronomia Lisbon, Portugal Lucie Parenicov DSM Biotechnology Centre, DSM Delft, The Netherlands Maria Teresa Pretel Escuela Politcnica Superior de Orihuela Universidad Miguel Hernndez Alicante, Spain Felix Romojaro Departamento Biologa del Estrs y Patologa Vegetal Centro de Edafologa y Biologa Aplicada del Segura-CSIC Espinardo, Murcia, Spain Johannes A. Roubos DSM Biotechnology Centre, DSM Delft, The Netherlands 94335.indb 12 3/31/10 4:29:08 PM 2010 Taylor and Francis Group, LLC 13. List of Contributors xiii Paloma Snchez-Bel Departamento Biologa del Estrs y Patologa Vegetal Centro de Edafologa y Biologa Aplicada del Segura-CSIC Espinardo, Murcia, Spain Luigi Servillo Dipartimento di Biochimica e Biofisica Seconda Universit degli Studi di Napoli Napoli, Italy Jun Song Agriculture and Agri-Food Canada Atlantic Food and Horticulture Research Centre Nova Scotia, Canada Marco A. van den Berg DSM Biotechnology Centre, DSM Delft, The Netherlands 94335.indb 13 3/31/10 4:29:08 PM 2010 Taylor and Francis Group, LLC 14. 94335.indb 14 3/31/10 4:29:08 PM 2010 Taylor and Francis Group, LLC 15. 1 onechapter Introduction to enzymes Alev Bayndrl 1.1 Nature of enzymes Enzymes are effective protein catalysts for biochemical reactions. The structural components of proteins are L--amino acids with the exception of glycine, which is not chiral. The four levels of protein structure are pri- mary, secondary, tertiary, and quaternary structures. Primary structure is related to the amino acid sequence. The amino group of one amino acid is joined to the carboxyl group of the next amino acid by covalent bonding, known as a peptide bond. The amino acid side-chain groups vary in terms of their properties such as polarity, charge, and size. The polar amino acid side groups tend to be on the outside of the protein where they interact with water, whereas the hydrophobic groups tend to be in the interior part of the protein. Secondary structure (-helix, -pleated sheet, and turns) is important for protein conformation. Right-handed -helix is a regular arrangement of the polypeptide backbone by hydrogen bonding between the carbonyl oxygen of one residue (i) and the nitrogenous proton of the other residue (i+4). -pleated sheet is a pleated structure composed of poly- peptide chains linked together through interamide hydrogen bonding between adjacent strands of the sheet. Tertiary structure refers to the three- dimensional structure of folded protein. Presence of disulfide bridges, hydrogen bonding, ionic bonding, and hydrophobic and van der Waals interactions maintain the protein conformation. Folding the protein brings Contents 1.1 Nature of Enzymes....................................................................................1 1.2 Enzyme Classification and Nomenclature.............................................2 1.3 Enzyme Kinetics........................................................................................3 1.4 Factors Affecting Enzyme Activity.........................................................7 1.5 Enzyme Inactivation............................................................................... 10 1.6 Enzymes in Fruit and Vegetable Processing........................................ 13 Abbreviations.................................................................................................... 16 References........................................................................................................... 16 94335.indb 1 3/31/10 4:29:09 PM 2010 Taylor and Francis Group, LLC 16. 2 Alev Bayndrl together amino acid side groups from different parts of the amino acid sequence of the polypeptide chain to form the enzyme active site that con- sists of a few amino acid residues and occupies a relatively small portion of the total enzyme volume. The rest of the enzyme is important for the three-dimensional integrity. The quaternary structure of a protein results from the association of two or more polypeptide chains (subunits). Specificity and catalytic power are two characteristics of an enzyme. Most enzymes can be extremely specific for their substrates and catalyze reactions under mild conditions by lowering the free energy require- ment of the transition state without altering the equilibrium condition. The enzyme specificity depends on the conformation of the active site. The enzyme-substrate binding is generally explained by lock-and-key model (conformational perfect fit) or induced fit model (enzyme confor- mation change such as closing around the substrate). The lock-and-key model has been modified due to the flexibility of enzymes in solution. The binding of the substrate to the enzyme results in a distortion of the substrate into the conformation of the transition state, and the enzyme itself also undergoes a change in conformation to fit the substrate. Many enzymes exhibit stereochemical specificity in that they catalyze the reac- tions of one conformation but not the other. Catalytic power is increased by use of binding energy, induced-fit, proximity effect, and stabilization of charges in hydrophobic environment. The catalytic activity of many enzymes depends on the presence of cofactor for catalytic activity. If the organic compound as cofactor is loosely attached to enzyme, it is called a coenzyme. It is called a prosthetic group when the organic compound attaches firmly to the enzyme by covalent bond. Metal ion activators such as Ca++, Cu++, Co++, Fe++, Fe+++, Mn++, Mg++, Mo+++,and Zn++ can be cofactors. An enzyme without its cofactor is called an apoenzyme. An enzyme with a cofactor is referred as a haloenzyme. Enzymes catalyze the reactions by covalent catalysis or general acid/base catalysis. 1.2 Enzyme classification and nomenclature Enzymes are classified into six groups (Table1.1) according to the reaction catalyzed and denoted by an EC (Enzyme Commission) number. The first, second, and thirdfourth digits of these numbers show class of the enzyme, type of the bond involved in the reaction, and specificity of the bond, respectively. Systematic nomenclature is the addition of the suffix -ase to the enzyme-catalyzed reaction with the name of the substrate. For example, naringinase, and -L-Rhamnoside rhamnohydrolase are trivial and system- atic names of the enzyme numbered as EC 3.2.1.40, respectively. Some of the enzyme-related databases are IUBMB, International Union of Biochemistry and Molecular Biology enzyme no menclature (www.chem.qmul. ac.uk/ iubmb/enzyme/); BRENDA, comprehensive enzyme information system 94335.indb 2 3/31/10 4:29:09 PM 2010 Taylor and Francis Group, LLC 17. Chapter one: Introduction to enzymes 3 (www.brenda-enzymes.org); the ExPASy, Expert Protein Analysis System enzymenomenclature(www.expasy.org/enzyme/);andEBIPDB,European Bioinformatics InstituteProtein Data Bank enzyme structures database (www.ebi.ac.uk/thornton-srv/databases/enzymes). 1.3 Enzyme kinetics Besides the quasi-steady-state kinetics (Briggs and Haldane approach), the rate of enzyme catalyzed reactions is generally modeled by the Michaelis-Menten approach. For a simple enzymatic reaction, bind- ing of substrate (S) with free enzyme (E) is followed by an irreversible Table1.1 Classification of Enzymes EC Class Reaction Catalyzed Examples EC1: Oxidoreductases A B A B + + Peroxidase Catalase Polyphenol oxidase Lipoxygenase Ascorbic acid oxidase Glucose oxidase Alcohol dehydrogenase EC2: Transferases AB C A BC+ + Amylosucrase Dextransucrase Transglutaminase EC3: Hydrolases AB H O AH BOH+ +2 Invertase Chlorophyllase Amylase Cellulose Polygalacturonase Lipase Galactosidase Thermolysin EC4: Lyases A B A B X Y X Y = + Pectin lyase Phenylalanine ammonia lyase Cysteine sulfoxide lyase Hydroperoxide lyase EC5: Isomerase A B A B X Y Y X Glucose isomerase Carotenoid isomerase EC6: Ligases A B AB+ Hydroxycinnamate CoA ligase 94335.indb 3 3/31/10 4:29:13 PM 2010 Taylor and Francis Group, LLC 18. 4 Alev Bayndrl breakdown of enzyme-substrate complex (ES) to free enzyme and prod- uct (P). The substrate binding with E is assumed to be very fast relative to the breakdown of ES complex to E and P. Therefore, the substrate binding is assumed to be at equilibrium as shown in the following reac- tion scheme: E S ES E P k k k + + 1 1 2 (1.1) The Michaelis-Menten approach concerns the initial reaction rate where there is very little product formation. It is impossible to know the enzyme concentration in enzyme preparations. Therefore, the amount of the enzyme is given as units of activity per amount of sample. One inter- national enzyme unit is the amount of enzyme that produces 1 micro- mole of product per minute. According to the applied enzyme activity assay, the enzyme unit definition must be clearly stated in research or application. The total enzyme amount (Eo) equals the sum of the amount of E and ES complex. In terms of amounts, it can be represented as follows: C C CE E ESo = + (1.2) The dissociation constant (Km), which is also called the Michaelis- Menten constant, is a measure of the affinity of enzyme for substrate: K k k C C Cm E S ES = =1 1 (1.3) If the enzyme has high affinity for the substrate, then the reaction will occur faster and Km has a lower value. High Km value means less affinity. Km varies considerably from one enzyme to another and also with differ- ent substrates for the same enzyme. For these elementary reactions 1.1, the initial reaction rate or reaction velocity (v) is expressed as v k CES= 2 (1.4) If the enzyme is stable during the reaction, the maximum initial reac- tion rate (vmax) corresponds to v k CEomax = 2 (1.5) 94335.indb 4 3/31/10 4:29:16 PM 2010 Taylor and Francis Group, LLC 19. Chapter one: Introduction to enzymes 5 If the initial concentration of substrate (So) is very high during the reaction (CSo >> CEo), the concentration of substrate remains constant dur- ing the initial period of reaction (CSo CS). Combining Equations 1.21.5, the Michaelis-Menten equation is obtained as v v C K C S m S = + max (1.6) As an example, kinetic properties of polygalacturonase assayed in different commercial enzyme preparations were studied and the reac- tions in all samples followed MichaelisMenten kinetics (Ortega et al., 2004) Michaelis-Menten expression can be simplified as follows: zero order ression forCexp : maxv v Ks m= >> (1.7) first order ression for Cexp : max v v K C K m S s m=