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Copyright Statement This Copy of the Thesis Has Been Supplied on The University of Plymouth PEARL https://pearl.plymouth.ac.uk 04 University of Plymouth Research Theses 01 Research Theses Main Collection 2017 Lactic acid bacteria as bio-preservatives in bakery Role of sourdough systems in the quality, safety and shelf life of bread Koy, Rebaz http://hdl.handle.net/10026.1/9828 University of Plymouth All content in PEARL is protected by copyright law. Author manuscripts are made available in accordance with publisher policies. Please cite only the published version using the details provided on the item record or document. In the absence of an open licence (e.g. Creative Commons), permissions for further reuse of content should be sought from the publisher or author. Copyright Statement This copy of the thesis has been supplied on the condition that anyone who consults it is understood to recognise that its copyright rests with its author and that no quotation from the thesis and no information derived from it may be published without the author’s prior consent. Lactic acid bacteria as bio-preservatives in bakery – Role of sourdough systems in the quality, safety and shelf life of bread by Rebaz Aswad Mirza Koy A thesis submitted to Plymouth University in partial fulfilment for the degree of DOCTOR OF PHILOSOPHY School of Biological and Marine Sciences Faculty of Science and Engineering August 2017 Lactic acid bacteria as bio-preservatives in bakery – Role of sourdough systems in the quality, safety and shelf life of bread Rebaz Aswad Mirza Koy Abstract Microbial contamination and survival during storage of bread are a cause of both health concerns and economic losses. Traditional fermentation systems were studied as sources of lactic acid bacteria (LAB) with antagonistic potential against foodborne pathogens and spoilage organisms, with the aim to improve the safety and shelf life of bakery products. The antagonistic activity of four types of buttermilk (BM) products fermented with Lactococcus lactis subsp. lactis was evaluated against a number of pathogenic bacteria to select the best fermented-BM for application as bio-preservatives in bread crumpets, showing up to 9 µg/ml of nisin equivalent antimicrobial activity. These food ingredients could be suitable to be used in crumpet formulations, BM fermented with Lc. lactis subsp. lactis and nisin influenced the quality and shelf life of crumpets; the pH value and firmness of products with fermented BM was lower and the acidity and springiness was higher than for unfermented BM treatment and control withouth additive. The nisin and fermented BM treatment had beneficial effects on the pore size and colour in comparison with the control, and improved microbial shelf life by 2 days. Commercial and traditional sourdough and bread samples (n=18) were collected to assess the diversity of LAB strains and potential properties when applied to dough and bread. DGGE followed by sequencing showed that Lactobacillus was the predominant genus in the studied sourdoughs. Lb. plantarum and Lb. brevis strains accounted for 69% of the 32 isolates, out of which 10 were amylolytic and 12 had proteolytic activity. Most were also good acid producers after 24 h at 30°C. Some LAB strains presented a strong in vitro inhibitory activity against five indicator strains, showing potential as starter cultures to ferment sourdough. In subsequent experiments, the properties of 24 sourdoughs were evaluated, and one of them, fermented with Lb. plantarum (SIN3) yielded low pH value, high lactic acid production, and suitable microbial growth, and was selected for further bread making performance trials. The bread with fast fermentation and high sourdough concentration (FFHSD) had a lower pH, higher acidity and increased the quality attributes with significantly better shelf life comparing to the other treatments during the storage period. Sensory evaluation demonstrated that fast-fermented breads were more acceptable than the slow-fermented counterparts. Bread prepared with high level (18%) of sourdough fast-fermented with the selected culture (SIN3) had a good eating quality and shelf life. The approach of this study is likely to yield feasible improvements of the current methods of preparation of baking goods. I List of contents Abstract ............................................................................................................... I List of contents ................................................................................................... II List of tables ...................................................................................................... XI List of figures .................................................................................................. XIV List of abbreviations ........................................................................................ XVI Acknowledgments ........................................................................................ XVIII Dedication ....................................................................................................... XIX Author’s Declaration ....................................................................................... XXI II CHAPTER ONE ................................................................................................. 1 General introduction and literature review ..................................................... 1 1.1 Introduction ................................................................................................... 1 1.2 The concept of LAB ...................................................................................... 4 1.3 The concept of safety and fermented food ................................................... 6 1.4 Cereal microflora .......................................................................................... 8 1.5 Spoilage and pathogenic microorganisms associated with bread and bakery products ................................................................................................ 10 1.6 Chemical preservatives .............................................................................. 12 1.7 Use of LAB and their metabolites in the food industry as bio- preservatives .................................................................................................... 15 1.8 Antimicrobial compounds produced by LAB ............................................... 18 1.8.1 Organic acids (lactic acid and acetic acid) ........................................... 18 1.8.2 Hydrogen peroxide (H2O2) .................................................................... 19 1.8.3 Carbon Dioxide (CO2) ........................................................................... 20 1.8.4 Diacetyl................................................................................................. 20 1.8.5 Bacteriocins .......................................................................................... 21 1.8.5.1 Classification of bacteriocins .......................................................... 23 1.8.5.2 Structure of nisin and mode of action ............................................. 23 1.8.5.3 Usage of natural and commercial additives of nisin: (Nisin and its applications) .......................................................................................... 28 1.8.6 Production of exopolysaccharides (EPSs)............................................ 31 1.9 Safety concerns and shelf life of bakery products ...................................... 33 1.10 Overview of sourdough technology .......................................................... 38 1.10.1 LAB in sourdough fermentation and their Influence on sourdough ..... 41 1.10.2 Classification of sourdough ................................................................ 45 1.10.3 Use of sourdough in cereal products .................................................. 48 1.10.4 Proteolysis and starch hydrolysis ....................................................... 50 1.10.5 Effect of ingredients and processing on the sourdough flavour .......... 53 1.10.6 Beneficial effects of sourdough fermentation on bread quality ........... 55 1.11 The rationale ............................................................................................. 60 1.12 The aim and objectives of this study ......................................................... 63 III CHAPTER TWO ............................................................................................... 65 General materials and methods .................................................................... 65 2.1 Methods ...................................................................................................... 65 2.2 Bacterial strains .......................................................................................... 66 2.3 Culture media ............................................................................................. 66 2.3.1 Nutrient broth and agar ........................................................................ 66 2.3.2 Brain heart infusion agar (BHI) ............................................................. 67 2.3.3 Bacillus cereus selective agar base medium ........................................ 67 2.3.4 M17 ...................................................................................................... 67 2.3.5 Preparation of De Man, Rogosa Sharpe agar (MRS) ........................... 68 2.3.6 Potato Dextrose agar (PDA) ................................................................. 68 2.3.7 Yeast extract glucose chloramphenicol
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