THE EFFECTS OF CARROT CAROTENOIDS ON VISUAL FUNCTION IN LONG-HOUR COMPUTER USERS: A PILOT STUDY by MORGAN MURRAY A Thesis Submitted to the Faculty of Graduate Studies of the University of Manitoba in Partial Fulfillment of the Requirements for the Degree of MASTER OF SCIENCE Department of Human Nutritional Sciences University of Manitoba Winnipeg, Manitoba R3T 2N2 Copyright © 2014 Morgan Murray Abstract Although carotenoids are essential for visual function, their potential beneficial role in Computer Vision Syndrome (CVS) remains to be elucidated. By providing carrot powder rich in α- and β-carotene this study examined whether carrot carotenoids can influence retinal function in long-hour computer users. A double-blind, placebo- controlled, repeated measures pilot trial, consisting of male and female participants (n=19, ages 20-65) with CVS, were randomly assigned to two supplementation groups; control (15g cream of wheat powder) or carrot enriched (15g carrot powder, 33% of vitamin A RDA for adults) in the form of an isocaloric pudding and yogurt, every day for 4 weeks. Retinal function was assessed with the electroretinogram (ERG) at PRE (week 0) and POST (week 4) time points. Plasma oxidative stress markers, lipids, and carotenoid/retinoid levels were assessed using ELISA, auto-analyzer, and UPLC, respectively, at PRE, DURING (week 2), and POST. Self-perceived vision status was assessed using the Ocular Surface Disease Index questionnaire. Carrot supplementation marginally improved (P<0.1) photopic b-wave amplitudes, without reaching statistical significance, representing cone-driven phototransduction, in 75% of total retinas, indicating retinal sensitivity to dietary nutrients. Carrot supplementation significantly increased plasma retinol and β-carotene levels (P<0.02), however, were not associated with CVS symptom improvement. HDL cholesterol significantly increased, whilst LDL cholesterol exhibited a diet, lowering trend (P<0.09). Plasma F2-isoprostanes displayed a trending reduction (P<0.1) compared to the control group illustrating the anti-oxidant potential of the carrot. Improvements seen in cone-driven inner retinal responses, along with increased plasma carotenoid/retinoid levels and beneficial lipid and oxidative stress changes, indicate that minimal supplementation of carotenoids at 33% of the vitamin A RDA by carrot powder can be recommended as a novel nutritional therapy for healthy, chronic computer users. A larger sample size test is warranted to obtain robust results. 2 Acknowledgements I would like to express my gratitude towards my supervisor Dr. Miyoung Suh for her utmost support, expertise and guidance throughout my research. A special acknowledgment also extends to my committee Dr. B. Albensi, Dr. M. Aliani, and Dr. L. Bellan for their advice and knowledge. Many thanks to my lab members for their generous assistance and for making my time a memorable experience. Additional thanks to Dennis Labossiere and Haifeng Yang for their skilled assistance in the analytical component of the project. This research would not have been possible without the generous financial support of the University of Manitoba (Manitoba Graduate Scholarship), which provided me with studentships for the duration of my degree. I would also like to thank Manitoba Agriculture, Food and Rural Initiatives – MB Agri-Health Research Network (MAHRN) and Food Product Development Centre for supporting the research. Finally, my deepest love and thanks extend to my parents, and my boyfriend, Marshall, for their endless support and encouragement. 3 Table of Contents List of Tables ............................................................................................................................... 7 List of Figures .............................................................................................................................. 7 List of Abbreviations ................................................................................................................... 9 Chapter I: INTRODUCTION ........................................................................................................ 12 Carotenoid and vitamin A metabolism ...................................................................................... 13 Classifications ........................................................................................................................ 13 Absorption and metabolism ................................................................................................... 13 Vitamin A deficiency ................................................................................................................. 20 Vitamin A toxicity ..................................................................................................................... 23 Dietary recommendations .......................................................................................................... 24 Retina anatomy .......................................................................................................................... 25 Retinal pigment epithelium ........................................................................................................ 26 Electroretinogram ...................................................................................................................... 28 Carotenoids and Vitamin A in visual function........................................................................... 31 Rod visual cycle ..................................................................................................................... 31 Cone visual cycle ................................................................................................................... 33 Other carotenoids in visual function ...................................................................................... 34 Carotenoids in chronic disease ................................................................................................... 36 Carotenoid supplementation studies .......................................................................................... 37 Computer Vision Syndrome ...................................................................................................... 38 Symptoms .............................................................................................................................. 38 Prevalence .............................................................................................................................. 39 Treatment ............................................................................................................................... 41 Carrot as a source of vitamin A and carotenoids ....................................................................... 43 Carrot as a health food ............................................................................................................... 45 4 Chapter II. RESEARCH PLAN ..................................................................................................... 47 Rationale .................................................................................................................................... 47 Hypothesis ................................................................................................................................. 49 Objectives .................................................................................................................................. 49 Chapter III. EXPERIMENTAL DESIGN AND METHODS ........................................................ 50 Experimental Design ...................................................................................................................... 50 Study Design .............................................................................................................................. 50 Participants ................................................................................................................................. 51 Diet Supplements ....................................................................................................................... 52 Questionnaires ........................................................................................................................... 53 Lifestyle ................................................................................................................................. 53 Ocular Surface Disease Index Questionnaire ......................................................................... 54 Experimental Methods ................................................................................................................... 54 Blood Parameters ....................................................................................................................... 54 Plasma Glucose .......................................................................................................................... 54 Plasma Lipids ............................................................................................................................. 55 Electroretinography.................................................................................................................... 55 Carrot powder carotenoid composition and stability measurement ........................................... 57 Enzyme-linked immunosorbent assay of oxidative stress markers............................................ 60 Statistical Analysis ....................................................................................................................