Biotechnology of Natural Products (eBook)

Dr. Wilfried Schwab is an Associate Professor of Biotechnology of Natural Products at Technische Universität München in Freising, Germany. Dr. Bernd Markus Lange is an Associate Professor at the Institute of Biological Chemistry, M.J. Murdock Metabolomics Laboratory, at Washington State University in Pullman, Washington. Dr. Matthias Wüst is an Associate Professor in the Institute of Nutritional and Food Sciences at Universität Bonn in Bonn, Germany. 

Preface 5
Contents 7
Contributors 9
Part I: Biotechnological Production of Selected Natural Products 12
1: Vanilla: The Most Popular Flavour 13
1.1 Introduction 13
1.2 Vanilla Orchids, Vanilla Flowers and the Pod 14
1.3 When the Green Becomes Black – Vanilla Pod Curing 17
1.4 Vanillin Is the Key Flavour Compound of the Complex Vanilla Extract 18
1.5 How Does the Vanilla Plant Form Vanillin? 20
1.6 Flavour Synthesis by Brewing – Bioengineering of Vanillin Biosynthesis 21
1.7 Biotechnology-Based Production of Vanillin from Eugenol, Iso-Eugenol, Ferulic Acid and Glucose 24
1.8 Future Perspectives and Final Remarks 29
References 30
2: Rosmarinic Acid and Related Metabolites 35
2.1 Occurrence and Structures of Rosmarinic Acid and Related Metabolites 36
2.2 Biosynthetic Pathway of Rosmarinic Acid 42
2.3 Production of RA in Untransformed Aseptic In Vitro Cultures 44
2.3.1 Species from the Family Lamiaceae 44
2.3.2 Species of the Family Boraginaceae 50
2.3.3 Non-vascular Plant Species 52
2.4 Production of Rosmarinic Acid in Hairy Roots 52
2.4.1 Hairy Roots of Lamiaceae Species 52
2.4.2 Hairy Roots of Boraginaceae Species 57
2.5 Production of Rosmarinic Acid and Related Caffeic Acid Esters in Microorganisms 58
2.6 In Vitro Formation of Non-natural Hydroxycinnamic Acid Esters and Amides by “Rosmarinic Acid Synthase” 60
2.7 Conclusion and Outlook 61
References 61
3: Bioproduction of Resveratrol 71
3.1 Introduction 71
3.2 Biosynthesis of Resveratrol and Its Analogs 72
3.3 Bioproduction of Resveratrol in Microorganisms 73
3.3.1 Yeast 74
3.3.2 E. coli 78
3.3.3 Other Bacteria 79
3.4 Bioproduction of Resveratrol Analogs and Derivatives 80
3.5 Strategies in Metabolic Engineering of Resveratrol Production 81
3.5.1 Pathway Engineering to Increase Precursor Supply 82
3.5.2 Protein Engineering 83
3.6 Conclusion and Perspective 84
References 85
4: Anthocyanin Production in Engineered Microorganisms 90
4.1 Anthocyanins and Their Industrial Applications 91
4.1.1 Pharmaceutical Applications 91
4.1.2 Food Colorants 92
4.1.3 Cosmetic Industry 92
4.1.4 Other Fields 93
4.2 Plant-Based Anthocyanin Production 93
4.2.1 Extraction from Plants 94
4.2.2 Anthocyanin Production from Suspension Cell Culture 94
4.3 Anthocyanin Production in Microorganisms 96
4.3.1 Engineering of Pathway Enzymes 97
4.3.2 Supply of Cofactors and Cosubstrates 99
4.3.3 Engineering Anthocyanin Secretion 100
4.3.4 Optimization of the Production Process 100
4.4 Conclusions and Future Perspectives 101
References 102
5: Microbial Synthesis of Plant Alkaloids 107
5.1 Introduction 109
5.2 Reconstitution of MIA Biosynthetic Pathways 114
5.2.1 Precursor Pathways 115
5.2.2 Strictosidine Formation 116
5.2.3 Downstream Derivatization 117
5.3 Reconstitution of BIA Biosynthetic Pathways 118
5.3.1 Upstream BIA Pathways 118
5.3.2 Microbial Production of Morphinan Alkaloids 121
5.3.3 Microbial Production of Protoberberine, Benzophenanthridine, and Phthalideisoquinoline Alkaloids 122
5.3.3.1 Synthesis of Dihydrosanguinarine and Sanguinarine in Yeast 122
5.3.3.2 Noscapine Synthesis in Yeast 123
5.4 Microbial Engineering Challenges 124
5.4.1 Functional Expression of Cytochromes P450 (CYPs) 124
5.4.2 Taming Enzyme Promiscuity 125
5.4.3 Limiting Efflux of Intermediates 127
5.5 Optimization Strategies 127
5.5.1 Genetic and Pathway Engineering Techniques 127
5.5.1.1 Building Combinatorial Enzyme Libraries 128
5.5.1.2 Pathway Assembly 128
5.5.1.3 Tuning Gene Expression 129
5.5.2 Cultivation Methods 130
5.6 Conclusions and Future Directions 131
References 132
6: Caffeine 139
6.1 Introduction 139
6.2 Distribution of Caffeine in Plants 139
6.3 Biosynthesis of Caffeine from Xanthosine 141
6.4 The Caffeine Synthase Gene Family in Plants 145
6.5 Biotechnological Production of Caffeine by Genetic Engineering 148
References 148
7: Taxol® Biosynthesis and Production: From Forests to Fermenters 152
7.1 Introduction 152
7.2 The Biosynthesis of Taxol 155
7.3 Taxol Production by Endophytes 161
7.4 Taxol Production by Plant Cell Cultures 165
7.5 Microbial Biotechnology for the Production of Taxol 176
7.6 Concluding Remarks 179
References 180
Part II: Technologies for Metabolic, Enzyme and Process Engineering 193
8: Commercial-Scale Tissue Culture for the Production of Plant Natural Products: Successes, Failures and Outlook 194
8.1 Introduction 194
8.2 Commercial Products from Plant Tissue Culture 197
8.3 Considerations for Commercial Targets 201
8.4 Future Opportunities 217
References 219
9: Tailoring Natural Products with Glycosyltransferases 224
9.1 Introduction 225
9.2 The Significance of Glycosylation in Plants 227
9.2.1 Glycosylation Increases Solubility 227
9.2.2 Glycosylation Increases Stability 227
9.2.3 Glycosylation Controls Sequestration/Compartmentalization 228
9.2.4 Glycosylation Affects Bioactivity and -Availability 229
9.2.5 Glycosylation Reduces Toxicity 230
9.2.6 Glycosylation Affects Perception 231
9.3 Glucoside/Glucose Ester Synthesis 231
9.4 Family 1 Plant Glycosyltransferases 233
9.5 Substrates of Family 1 Plant Glycosyltransferases 234
9.5.1 Secondary Metabolites 234
9.5.1.1 Phenylpropanoids 235
9.5.1.2 Flavonoids, Anthocyanins 236
9.5.1.3 Dihydrochalcones, Acylphloroglucinol, Stilbenes, Curcumin 236
9.5.1.4 Terpenoids 238
9.5.2 Plant Hormones 239
9.5.3 Miscellaneous Substrates (Alkaloids, Benzoxazinoids, Furanones, and Xenobiotics) 239
9.6 Glucoside Production by Whole Cell Biocatalysts 240
9.6.1 Production System 240
9.6.2 Types of Whole-Cell Biocatalysts 247
9.6.3 Process Optimization 247
9.6.3.1 Vector Conveyed Process Optimization 248
9.6.3.2 UGT Optimization 248
9.6.3.3 Host Genome Optimization – Metabolic Engineering 249
9.7 Recent Applications of Glycosyltransferases for Production of Small Molecule Glycosides 252
9.7.1 Glycosyltransferases and Glycosylation in Product for Consumer Consumption 252
9.7.2 Glycorandomization 254
9.7.3 Glycosyltransferases and Their Role in Cancer Therapies 254
9.8 Conclusions and Future Prospects 255
References 256
Part III: Analytical and Legal Aspects 269
10: Authenticity Control of Natural Products by Stable Isotope Ratio Analysis 270
10.1 Introduction 270
10.2 Isotope Ratio Mass Spectrometry – IRMS 272
10.2.1 Notations in IRMS 272
10.2.2 Elemental Analyzer-IRMS (EA-IRMS) and Gaschromatography-IRMS (GC-IRMS) 273
10.2.3 Liquid Chromatography-IRMS (LC-IRMS) 275
10.2.4 Site-Specific Natural Isotope Fractionation-Nuclear Magnetic Resonance Spectroscopy (SNIF-NMR) 275
10.3 Practical Applications of Stable Isotope Analysis on Miscellaneous Natural Products 276
10.3.1 Phenylpropanoids (Vanillin) 276
10.3.2 Terpenes (Monoterpenes and Tetraterpenes) 278
10.3.3 Polyphenols (Resveratrol) 279
10.3.4 Alkaloids (Caffeine) 280
10.4 Conclusions 280
References 280
11: Natural or Synthetic? The Legal Framework in the EU for the Production of Natural Flavouring Ingredients 283
11.1 Introduction 283
11.2 General Conditions 284
11.3 Scope of the Regulation 284
11.4 Definitions 285
11.4.1 Definitions of Flavourings in General 285
11.4.2 Definition of Natural Flavouring Substance 286
11.4.3 Definitions of Flavouring Preparations 290
11.5 Processes for the Production of Natural Flavouring Ingredients 294
11.5.1 EFFA Considerations on the Permitted Order of the Various Processes 294
11.5.2 Biotechnology for the Production of Natural Flavouring Ingredients 295
11.5.3 Further Considerations on the Production of Natural Flavouring Substances and Flavouring Preparations According to the EFFA Guidance Document 296
11.6 Analytical Methods to Assess Authenticity 297
11.7 Labelling of Flavourings (B2B) 298
11.7.1 General Labelling Requirements 298
11.7.2 Labelling of Natural Flavourings 299
11.7.3 Considerations on Different Interpretations of the 95/5-­Rule (Art. 16(4)) 302
11.8 Safety Evaluation of Flavourings and Their Inclusion in the EU Union List 303
References 306
Index 308