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dc.contributor.authorMa, Yanwei
dc.date.accessioned2019-08-09
dc.date.available2019-08-09
dc.date.issued2018-10-05
dc.identifier.urihttp://hdl.handle.net/2123/20877
dc.description.abstractVitamin K has emerged as a multi-functional vitamin that is essential in a diverse range of physiological effects in the human body. Among all the vitamin K family members, menaquinone-7 (vitamin K2) has gained the most popularity for the prevention and treatment of osteoporosis and atherosclerosis. The aim of this study was to develop a strategy to produce high level of MK-7 via fermentation by Bacillus subtilis for applications in the food and pharmaceutical industries. A series of screening and optimisation steps were carried out to develop a viable MK-7 formula for fortified food application. The highest titre of 30 mg/L MK-7 was achieved by using 10 w/v % soy protein and 5 w/v% glycerol supplemented with 0.5 w/v % yeast extract and 0.6 % K2HPO4.The major problem of foam formation in stirred fermenter was tackled by using a novel strategy of controlled dual feeding of glycerol and soy protein simultaneously. The MK-7 concentration in the fermentation was increased by four fold, from 30 mg/L in a 100 mL shake flask to 112 mg/L in a 2 L fed-batch bioreactor. Metabolic engineering was another approach undertaken to increase MK-7 production. 12 engineered B. subtilis 168 strains were constructed by individually overexpressing or co-overexpressing the rate limiting MEP pathway genes dxs, dxr, idi and the downstream gene menA in different orders. An ordered overexpression of menA-dxs-dxr-idi produced the highest titre of MK-7 at 50 mg/L in 50 mL SYG media in shake flasks. This improvement was 11-fold higher than the B. subtilis 168 control strain (4.5 mg/L) and 2.2-fold higher than the best-performing B. subtilis natto strain (22 mg/L). In summary, the MK-7 production was significantly increased by process optimisation and metabolic engineering strategies in this study. This study also implies the potential of using B. subtilis as the cellular factory for production of other industrially important MEP pathway derived molecules.en_AU
dc.publisherUniversity of Sydneyen_AU
dc.publisherFaculty of Engineeringen_AU
dc.publisherSchool of Chemical and Biomolecular Engineeringen_AU
dc.rightsThe author retains copyright of this thesis. It may only be used for the purposes of research and study. It must not be used for any other purposes and may not be transmitted or shared with others without prior permission.en_AU
dc.subjectVitamin Ken_AU
dc.subjectfunctional fooden_AU
dc.subjectBacillus subtilisen_AU
dc.subjectnattoen_AU
dc.subjectMEP pathwayen_AU
dc.subjectbiofilmen_AU
dc.titleProduction of Menaquinone-7 for Food and Pharmaceutical Applicationsen_AU
dc.typePhD Doctorateen_AU
dc.type.pubtypeDoctor of Philosophy Ph.D.en_AU
dc.description.disclaimerAccess is restricted to staff and students of the University of Sydney . UniKey credentials are required. Non university access may be obtained by visiting the University of Sydney Library.en_AU


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