الجمهورية الجزائرية الديمقراطية الشعبية وزارة التعليم العالي و البحث العلمي People’s Democratic Republic of Algeria Ministry of Higher Education and Scientific Research جامعة محمد الصديق بن يحي - جيجل University Mohammed Seddik Benyahia - Jijel كلية علوم الطبيعة و الحياة Faculty of Nature and Life Sciences قسم المكروبيولوجيا التطبيقية وعلوم التغذية Department of Applied Microbiology and Food Sciences Thesis Realized by Samiya AMIRA In Fulfillment of the Requirements for the Degree of Doctor of Sciences In Biology Option: Microbial Biotechnology Effect of microencapsulation on the viability of probiotic bacteria under storage and simulated gastrointestinal conditions Under the Supervision of: Pr. Mohammed SIFOUR Examiners committee Chairman: Pr. Tayeb IDOUI University Mohamed Seddik Benyahia-Jijel Supervisor: Pr. Mohammed SIFOUR University Mohamed Seddik Benyahia-Jijel Examiners: Dr. Hani BELHADJ University of Farhat Abbas-Setif 01 Dr. Abdelhakim AOUF University of Farhat Abbas-Setif 01 Dr. Abdelhafid BOUBENDIR University of Abdelhafid Boussouf -Mila Academic year: 2019/2020 Serial number: Acknowledgments Firstly, I thank Allah for all things given for me, I would like to express my sincere gratitude to my advisor Prof. Mohammed SIFOUR for the continuous support of my doctorate thesis and related research, for his patience, motivation, and immense knowledge. His guidance helped me in all the time of research and writing of this thesis. I could not have imagined having a better advisor for my doctorate study. A very special gratitude goes out to Prof. Houria OULED HADDAR and Prof. Tayeb IDOUI for helping me during the realization of this work. Besides my advisor, I would like to thank the members of thesis committee, the chairman Prof. Tayeb IDOUI, and the examiners Dr. Hani BELHADJ, Dr. Abdelhakim AOUF and Dr. Abdehafid BOUBENDIR for their acceptance to evaluate this work. I am grateful to my mother, who has provided me through moral and emotional support in my life. I am also grateful to my father, brother, sisters and friends who have supported me along the way. With a special mention to Dr. Asma Cherbal, for her patience, her help in material provided and thesis redaction. I acknowledge also, Ms. Saliha Hirache and Ms. Moufida Bensam for their emotional support. Another person to whom I address my special thanks, for his help in the thesis manuscript, thank you Ahmed. Big thanks for Ms. Sawsen Hadef, Mr. Tarek Khennouf and Mr.Mohammed Taher Boubezari. I am grateful to Ministry of Higher Education and Scientific Research to support this research. My sincere thanks also go to Pr. Gianluigi Mauriello, from the University of Federico II, Naples, Italy, who provided me an opportunity to join his laboratory team: Dr. Diamante Maresca, and Dr. Annachiara De prisco, who gave me the opportunity to access to the laboratory and research facilities. I am also grateful to the staff of Laboratory of Applied Microbiology and Laboratory of Molecular Toxicology at the University Mohamed Seddik Benyahia, Jijel. And finally, I thank everyone who contributed in the realization of this thesis. Thanks for all your encouragement ii List of content Acknowledgements ....................................................................................................................... ii List of abbreviations...................................................................................................................... iii List of figures ................................................................................................................................ iv List of tables .................................................................................................................................. vi Introduction ................................................................................................................................... 1 I. Literature review I.1. Lactic acid bacteria and probiotics ......................................................................................... 3 I.1.1. Probiotics ............................................................................................................................. 3 I.1.2. Lactic acid bacteria .............................................................................................................. 3 I.1.3. Screening and selection of probiotics .................................................................................. 5 I.1.4. Health benefits and mechanisms of action of LAB ............................................................ 6 I.2. Klila ........................................................................................................................................ 8 I.3. Microencapsulation ................................................................................................................. 11 I.3.1. Definition ............................................................................................................................. 11 I.3.2. Encapsulating materials ....................................................................................................... 12 I.3.2.1. Alginate............................................................................................................................. 12 I.3.2.2. Gellen gum........................................................................................................................ 12 I.3.2.3. Xanthan gum ..................................................................................................................... 13 I.3.2.4. Chitosan ............................................................................................................................ 13 I.3.2.5. Cellulose Acetate Phtalate (CAP) ..................................................................................... 13 I.3.2.6. Starch ................................................................................................................................ 13 I.3.2.7. K-carrageenan ................................................................................................................... 14 I.3.2.8. Gum Arabic (Acacia gum) ................................................................................................ 14 I.3.2.9. Locust bean gum .............................................................................................................. 15 I.3.2.10. Gelatin............................................................................................................................. 15 I.3.2.11.Whey and Milk proteins ................................................................................................. 15 I.3.2.12.Pectin ............................................................................................................................... 16 I.3.2.13. Chickpea protein ............................................................................................................. 16 I.3.3. Encapsulating techniques ..................................................................................................... 16 I.3.3.1. Extrusion ........................................................................................................................... 16 I.3.3.2. Emulsion ........................................................................................................................... 17 I.3.3.3. Spray drying...................................................................................................................... 18 I.3.3.4. Coacervation ..................................................................................................................... 19 I.3.3.5. Spray chilling/cooling/congealing .................................................................................... 19 I.3.3.6. Freeze-drying .................................................................................................................... 19 I.3.4. Factors affecting on microencapsulation effectiveness ....................................................... 19 I.3.5. Applications of microencapsulation .................................................................................... 20 I.3.6. Probiotics in food products .................................................................................................. 20 I.3.6.1. Dairy products ................................................................................................................. 20 I.3.7.1.1. Yoghurts ........................................................................................................................ 21 I.3.6.1.2. Cheese ............................................................................................................................ 21 I.3.6.1.3. Ice cream and frozen dairy desserts ............................................................................... 21 I.3.6.2. Non-dairy products ........................................................................................................... 21 I.3.6.3. Other food carriers ............................................................................................................ 22 II. Materials and methods II.1. Bacterial isolates ....................................................................................................................... 23 II.2. Media and chemicals ................................................................................................................. 23 II.3. Isolation and purification of lactic acid bacteria