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Leung Yiu Fai Ivan. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1997. / Includes bibliographical references (leaves 41-46). / Acknowledgements --- p.ii / Table of contents --- p.iii / List of tables --- p.vi / List of figures --- p.vii / List of Abbreviations --- p.xii / Abstract --- p.xiii / Chapter I. --- INTRODUCTION --- p.1 / Chapter A. --- Current knowledge on carotenoids --- p.1 / Chapter 1. --- Chemistry --- p.1 / Chapter 2. --- Occurrence --- p.3 / Chapter 3. --- Metabolism --- p.6 / Chapter 4. --- Biological function --- p.8 / Chapter B. --- Statement of the problem --- p.10 / Chapter II. --- MATERIAL AND METHODS --- p.11 / Chapter A. --- Materials --- p.11 / Chapter 1. --- Animals --- p.11 / Chapter 2. --- Human samples --- p.11 / Chapter 3. --- Chemicals --- p.12 / Chapter 4. --- HPLC Apparatus --- p.12 / Chapter B. --- Methods --- p.14 / Chapter 1. --- Animal specimens --- p.14 / Chapter (a) --- Carotenoid supplement --- p.14 / Chapter (b) --- Tissue preparation --- p.14 / Chapter 2. --- Human specimens --- p.15 / Chapter 3. --- Extraction of lipid component --- p.15 / Chapter (a) --- Retina --- p.15 / Chapter (b) --- Serum or subretinal fluid --- p.16 / Chapter (c) --- Liver --- p.16 / Chapter 4. --- Analytical methods --- p.17 / Chapter (a) --- Isocratic elution --- p.17 / Chapter (b) --- Gradient elution --- p.17 / Chapter III. --- RESULTS --- p.18 / Chapter A. --- Selection of chromatographic method for carotenoid analysis --- p.18 / Chapter 1. --- Effect of dioxane concentration on the retention time of carotenoids --- p.18 / Chapter (a) --- Gradient elution --- p.18 / Chapter (b) --- Isocratic elution --- p.18 / Chapter 2. --- Chromatograms of carotenoids and retinoids of a selected method for routine analysis --- p.19 / Chapter (a) --- Carotenoids --- p.20 / Chapter (b) --- Retinoids --- p.21 / Chapter B. --- Application of the selected method to study carotenoid in human and rats --- p.22 / Chapter 1. --- Study in human serum and subretinal fluid --- p.22 / Chapter (a) --- Serum --- p.22 / Chapter (b) --- Subretinal fluid --- p.22 / Chapter 2. --- Study in rats tissues --- p.24 / Chapter (a) --- Liver --- p.24 / Chapter (b) --- Serum --- p.26 / Chapter (c) --- Retina --- p.28 / Chapter C. --- Influence of dietary carotenoids on retinol concentration in rats --- p.31 / Chapter 1 --- Serum --- p.31 / Chapter 2 --- Retina --- p.31 / Chapter IV. --- DISCUSSION --- p.32 / Chapter A. --- Chromatographic analysis of carotenoids and retinoids --- p.32 / Chapter B. --- Carotenoid study in human and rats --- p.33 / Chapter 1. --- Carotenoids in human tissues --- p.33 / Chapter (a) --- Serum --- p.33 / Chapter (b) --- Subretinal fluid --- p.34 / Chapter 2. --- Dietary supplement of carotenoids to rats --- p.35 / Chapter (a) --- Choice of animal and types of carotenoids --- p.35 / Chapter (b) --- Carotenoids uptake into rats tissues --- p.36 / Chapter (c) --- Effect of dietary carotenoids on retinol concentration in rat tissues --- p.38 / Chapter V. --- CONCLUSION --- p.39 / Chapter VI. --- REFERENCES --- p.41 / Chapter VII. --- TABLES --- p.47 / Chapter VIII. --- FIGURES --- p.56

Thesis (M. Phil.)--University of Hong Kong, 2006. / Title proper from title frame. Also available in printed format.

Thesis (Ph. D.)--Massachusetts Institute of Technology, Dept. of Earth and Planetary Science, 1982. / Supervised by Robert B. Gagosian (W.H.O.I.). Vita. Includes bibliographies.

Thesis (Ph. D.)--Massachusetts Institute of Technology and Woods Hole Oceanographic Institution, 1982. / Includes bibliographical references.

Thesis (Ph. D.)--University of Alberta, 2009. / Title from pdf file main screen (viewed on Dec. 24, 2009). "A thesis submitted to the Faculty of Graduate Studies and Research in partial fulfillment of the requirements for the degree of Doctor of Philosophy in Microbiology and Cell Biotechnology, Department of Biological Sciences, University of Alberta." Includes bibliographical references.

Thesis (Ph. D.) -- Lincoln University, 2009. / Also available via the World Wide Web.

Thesis (M. Phil.)--University of Hong Kong, 2010. / Includes bibliographical references (leaves 97-109). Also available in print.

Astaxanthin (3,3'-dihydroxy-β-carotene-4,4'-dione) is a ketocarotenoid that is beneficial for human health due to its ability of boosting immune function and preventing tumor formation. The biosynthesis of astaxanthin is, however, limited only to a few organisms. The burgeoning demand for natural astaxanthin has attracted much recent interest in extending the carotenoid pathway of higher plants to astaxanthin by expressing a microbial β-carotene ketolase (BKT). One major challenge of engineering an astaxanthin pathway in plants is the low astaxanthin content achieved. Five green microalgae including Chlamydomonas reinhardtii, Chlorococcum sp., Neochloris wimmeri, Protosiphon botryoides and Scotiellopsis oocystiformis were selected with enhanced function for astaxanthin biosynthesis. The products of the BKT cDNAs from the algae are similar in sequence to the BKT from Haematococcus pluvialis (ca 70% amino acid identity). Based on an Escherichia coli system, the BKT enzymes were shown to exhibit various efficacies in converting zeaxanthin into astaxanthin with Chlamydomonas BKT exhibiting the highest conversion rate (ca 85%). To investigate if the function-enhanced Chlamdomonas BKT (CRBKT) has advantages over other algal BKTs in triggering astaxanthin biosynthesis in higher plants, the CrBKT, together with the BKTs from Chlorella zofingiensis (CzBKT) and H. pluvialis (HpBKT3) was expressed in Arabidopsis thaliana. Transgenic Arabidopsis expressing the CrBKT developed orange leaves which accumulated astaxanthin up to 2 mg g-1 dry weight. In contrast, the expression of CzBKT resulted in much lower content of astaxanthin (0.24 mg g-1 dry weight), whereas HpBKT3 was unable to mediate synthesis of astaxanthin in Arabidopsis. Similarly, overexpression of CrBKT in tobacco also resulted in the massive accumulation of astaxanthin in leaves (1.60 mg g-1 DW). Taken all together, it can be concluded that ketolating zeaxanthin efficiently is essential for high production of astaxanthin in transgenic plants. Tomato is an important food crop with high amounts of carotenoids in its fruit. To investigate if tomato fruit can serve as a bio-factory for astaxanthin production, the CrBKT was overexpressed in three genotypes of tomato. All transgenic tomato plants developed brown red leaves that accumulated canthaxanthin rather than astaxanthin as a major carotenoid, resulting from the poor catalytic activity of the endogenous BHY1 toward canthaxanthin. To overcome this problem, CrBKT and HpBHY, the best pair of genes catalyzing the formation of astaxanthin in β- carotene-producing E. coli, were coexpressed in tomato. Canthaxanthin was efficiently converted to astaxanthin, resulting in a massive accumulation of astaxanthin in leaf (3.12 mg g-1) and fruit (16.1 mg g-1) with enhanced total carotenoid capacities of 1.7-fold in leaf and 16.6-fold in fruit. Moreover, the over-production of astaxanthin in fruit enhanced its antioxidant capacity 3-5-fold and vitamin C 2-fold, although it did not affect growth and development. In summary, the Chlamydomonas BKT is proven to be superior to other sources of BKT/CrtW enzymes in triggering astaxanthin biosynthesis in plants. By coexpressing a pair of well-cooperating BKT and BHY genes, the transgenic B-type tomato could accumulate commercially attractive amounts of the high-value astaxanthin in its fruit. This study highlights the potential of higher plants to be engineered as cell factories for producing the high-value astaxanthin. / published_or_final_version / Biological Sciences / Doctoral / Doctor of Philosophy