Identification of the characteristic taste, aroma compounds and the corresponding precursors in pigmented rice wine Submitted for the degree of Doctor of Philosophy Department of Food and Nutritional Sciences School of Chemistry, Food and Pharmacy Sanchai Yotmanee September 2018 Declaration I confirm that the work presented in this thesis is my own and that the use of all the literature from other source has been properly and fully acknowledged. Sanchai Yotmanee September, 2018 Page | ii ABSTRACT Rice wine (Sake) is an alcoholic beverage which is produced using glutinous rice, fungi and yeast. It has a pale yellow to clear colour, umami taste and fruity aroma. Moreover, pigmented rice wine has been consumed in Asian countries for a long time, and the taste and flavour are distinctly different to Sake. Whereas Sake is produced from high polished rice and shows more clear colour and pleasant fruity aroma, whole pigmented rice is used to produce pigmented rice wine and its taste and aroma profiles are less known. In order to understand the differences in flavour and taste, commercial polished and unpolished pigmented rice wines were analysed for taste (sugars, organic acids, amino acids, phenolic acids, diketopiperazines (DKPs) and γ-glutamyl peptides) and aroma compounds. This study showed that compounds responsible for sweet, sour and metallic notes were found in commercial rice wines. These were attributed to glucose, acetic acid, cyclo(leucine-proline) and cyclo(isoleucine-proline). The aroma compounds in commercial rice wines were extracted using solid phase microextraction (SPME) and solid phase extraction (SPE) and analysed using gas chromatography-mass spectrometry (GC-MS) and GC-Olfactometry. Guaiacol and 4-vinylguaiacol were found to be the characteristic aroma compounds responsible for smoky-spicy note in commercial pigmented rice wine. The brewing process for pigmented rice wine was investigated to develop a constant and reproducible protocol for brewing of pigmented rice wine which can be used to investigate the formation of taste and aroma compounds. The pigmented rice was cooked by steaming or pressure cooking. The cooked rice was saccharified using Aspergillus oryzae, and fermented using Saccharomyces cerevisiae at 25 °C or 30 °C. This study showed that steaming was suitable for cooking rice. The brewing that was selected was 2 days for Page | iii saccharification and 9 days for fermentation at 30 °C. These conditions were based on obtaining higher concentration of sugars and ethanol. To study the formation of characteristic taste and aroma in pigmented rice wine, rice wines were brewed using unpolished (0%), 30% polished, 50% polished and 65% polished pigmented rice. They were analysed for taste and aroma compounds, and their precursors. The taste of lab-scale brewed rice wine was different from the commercial ones because astringent mouthfeel (gallic acid and protocatechuic acid) and umami were mainly found in lab-scale brewed rice wines, especially 0% RW. The bran increased the concentration of glutamic acid, phenolic acids and γ-glutamyl peptides (γ-glu-gly, γ-glu-his and γ-glu-tyr). The aroma compounds analysis also showed that guaiacol, 4-vinylguaiacol and vanillin were significantly and substantially increased in 0% RW. However, only guaiacol was found to be the characteristic aroma which contributed smoky-spicy note in 0% RW. This is consistent with sensory analysis which showed that a higher intensity of smoky-spicy note was found in 0% RW, compared to others. This study also confirmed that phenolic acids were derived from the pigmented rice bran by fungi and yeast during brewing. Vanillic acid was decarboxylated to form guaiacol. Moreover, guaiacol was also formed from unpolished pigmented rice by steaming. The sensory analysis confirmed that guaiacol was the key aroma compound which contributed smoky-spicy note in pigmented rice wine. Keywords: pigmented rice wine, characteristic aroma, smoky-spicy note, 4-vinylguaiacol, guaiacol Page | iv TABLE OF CONTENTS ABSTRACT iii TABLE OF CONTENTS v LIST OF TABLES x LIST OF FIGURES xiii ACKNOWLEDGEMENTS xvi CHAPTER 1: INTRODUCTION 1 1.1 Background .................................................................................................................................................. 1 1.2 Hypothesis ................................................................................................................................................... 3 1.3 Objectives of the study ............................................................................................................................ 3 1.4 Structure of the thesis ............................................................................................................................. 4 CHAPTER 2: LITERATURE REVIEW 6 2.1 Raw materials for the brewing of rice wine ..................................................................................... 6 2.1.1 Rice ......................................................................................................................................................... 6 2.1.2 Starter for fermentation of rice wine ...................................................................................... 10 2.2 Rice brewing process ............................................................................................................................ 12 2.2.1 Rice polishing ................................................................................................................................... 12 2.2.2 Rice cooking ..................................................................................................................................... 15 2.2.3 Fermentation of rice wine ........................................................................................................... 18 2.3 Aroma compounds in rice wine ........................................................................................................ 24 2.4 The formation of the aroma compounds during the brewing of rice wine ..................... 29 2.4.1 Higher alcohols, aldehydes and organic acids from amino acids ................................ 29 2.4.2 Esters ................................................................................................................................................... 30 2.4.3 Organic acids .................................................................................................................................... 33 Page | v 2.4.4 Volatile phenols .............................................................................................................................. 36 2.4.5 Lactones ............................................................................................................................................. 38 2.5 Taste compound in rice wine ............................................................................................................. 39 2.6 The formation of typical taste compounds in rice wine .......................................................... 43 2.6.1 Amino acids ...................................................................................................................................... 43 2.6.2 Phenolic acids .................................................................................................................................. 44 2.6.3 Diketopiperazines (DKPs) ........................................................................................................... 46 2.6.4 γ-glutamyl peptides ....................................................................................................................... 48 2.7 Concluding remarks .............................................................................................................................. 49 CHAPTER 3: TASTE AND AROMA COMPOUNDS IN RICE WINE: A COMPARISON OF POLISHED RICE WINE AND PIGMENTED RICE WINE 51 3.1 Introduction ............................................................................................................................................. 52 3.2 Materials and methods ........................................................................................................................ 53 3.2.1 Materials ............................................................................................................................................ 53 3.2.2 Chemicals .......................................................................................................................................... 53 3.2.3 Analysis of taste compounds in rice wine ............................................................................. 54 3.2.4 Aroma analysis for rice wines ..................................................................................................... 57 3.2.5 Statistical analysis ........................................................................................................................... 60 3.3 Results and discussions ........................................................................................................................ 60 3.3.1 Taste compounds in rice wine ................................................................................................... 60 3.3.2 The analysis of aroma in the commercial rice wines ......................................................... 73 3.4 Conclusion ...............................................................................................................................................