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University of Alberta

Library Release Form Name of Author: Vanessa Wing-Sze Lien Title of thesis: Supplementing Children with Arachidonic acid and

Docosahexaenoic acid improves Visual perception

Degree: Master of Science

Year this Degree Granted: 2005

Permission is hereby granted to the University of Alberta Library to reproduce single copies of this thesis and to lend or sell such copies for private, scholarly or scientific research purposes only.

The author reserves all other publication and other rights in association with the copyright in the thesis, and except as herein before provided, neither the thesis nor any substantial portion thereof may be printer or otherwise reproduced in any material form whatever without the author’s prior written permission.

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University of Alberta Supplementing Children with Arachidonic acid and Docosahexaenoic acid improves Visual perception by

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A thesis submitted to the Faculty of Graduate Studies and Research in partial fulfillment of the requirements for the degree of Master of Science

Nutrition and Metabolism

Department of Agricultural, Food and Nutritional Science

Edmonton, Alberta

Spring 2005

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Faculty of Graduate Studies and Research

The undersigned certify that they have read, and recommended to the Faculty of Graduate Studies and Research for acceptance, a thesis entitled Supplementing Children with Arachidonic acid and Docosahexaenoic acid Improves Visual Perception submitted by Vanessa Wing-Sze Lien in partial fulfillment of the requirements for the degree of Master of Science in Human Nutrition and Metabolism.

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Abstract

The objectives of this study were to 1) to determine dietary intakes of arachidonic acid (AA) and docosahexaenoic acid (DHA) in healthy children; 2) to supplement children (who consumed low intakes of DHA), with a nutritionally complete dietary Fala containing AA and DHA; 3) to evaluate AA and DHA fatty acid status in the blood and visual perception. Children, 4-7 years of age, living in central Alberta, Canada, who consumed low intakes of DHA were recruited in a controlled, double blind study and assigned randomly into two groups: the experimental group was provided with a formula containing AA (20-30 mg/day) and DHA (14-21 mg/day) (long chain polyunsaturated fatty acid group (LCP group)) and the control group was provided with the same formula without AA and DHA. There was a general trend toward a higher level of AA and DHA content (RBC phospholipid and plasma phospholipids) in LCP group compared to control group at 7 months. This study is the first to show that daily supplementation with AA and DHA for a period of 7 months improved visual perception in children who were

previously identified to have low dietary intakes of DHA.

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Table of Contents

Chapter 1.

CG TSTRS) AN CNSR TW ioe deco cc <a IF Intiedieronasaeeteer Metnod weol.io. Asacss. ek, ee DL, lomeoe” Ue [Ze DIN Ge ceo iC OMe DAMM ALPVEACI AS stu dins ns oc unitate sate ds ace ee ose ee oo aes 2

Ea Nomenclature.» ae aten. AS. ak. ELA, Wt oben cinevavanee es 2 P3 Essentialihatty Aicids-and:M etabolicsPathwaysit. # ORi8.....ccocecsacncenssacoare 3 PADIS tA BS OUI CES MERE REI ss rish slike dicisinn vsincdoabanesbedy dnaveesemebasbeenerweenseas 7% Reo WeE MO ANGIGs PALLY A CLOJUCTIY ALINGS smscja neiicceunnmenn tie cause eka meee eenmenc ee 7 DG Beem ese O A AULY A CLUS Bae ens cccs i at wired si Abe an neh omeee uae ean ee eae se 8 ie balance Between Omega o.and Omesa 3 Fatty Acids,...25 cue 9 1.8 Current Recommendations and Current Nutrient Intakes of Polyunsaturated BatiaAcidsanealy aomiater. Senplement com aes, AA. wel A Sot 7... 12 InStlgeNutrientdntakestoileAmand WN ARIA? SO Ie onc co ote dosloseucses 12 ASeaNuinent Intakes of AAs and, DEA ase oxssrcaoespnsecsctersievecctsoeniersou Soren casio 13 Mogg Mitte ROMAINE A a SSeMUalI tyre ages cies ieee dee hoe ae 16 1.9.1 Importance of AA and DHA: Retinal and Neuro Development......... 16 ih 9 22S YOLUNCSIS OlPAL PAT C NEAR BINED, «deca sha base eben ie eetteacanetseserane: i, poe eae A, INtAK esa Gio om a. deaue hat eine vere meee wee ete 18 1.10 Other nutrients of importance: Vitamin E, iron, and Vitamin A............... i COL MR anc ATTIRE PROM ieees ri Cicer arcs IA, eats cisco bins ee ERM otcat oaeertoiceeetncde 19 Clee ML ORs rt mk ah seme Nei i Ee, wide van Suis ne Saar en Het CUES fray 20 BORE 3S RAV AC ATA TEL AMUN ce ssh co Bee eee nad sank oy ico ute ur See mews te haan heim viesactes oe 22 PEL IMNCICTENCEAASIAUCOM ORO MARR a.c50 occ. .4csp ceckn ns nashesedudectenmmeusecpenuans 24

Chapter 2.

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Chapter 3.

Assessment of Docosahexaenoic Acid, Arachidonic Acid, Vitamin A, Vitamin E,

and iron Nutrienamnakesmu4 ~/.vear, old.G inidrens.2 aan.) tamee erences ase he aeeat eee 44 3:1) Introduction Meee Amare. tes Secs Prec rtey AMET nc on cee cine 44 3.2 Method sianmnregees on. 020s. Sembee A pce ee OY cece 46

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Chapter 4. Consuming a nutritionally complete supplement containing AA and DHA for 7 months improves visual perception in children 4-7 years of age.................0000008

4.1

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Food Assessment Method used to Assess AA and DHA Intakes........ 104

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Chapter 5.

Final sumimaneDiscussionand Conclusions: 1) ae) 2. eas Ot Beieete e - 119 Ome Cre Cea on TAL ye me eM MMMN nce et DiS ose \ ns ic Siar anes era wae ack regan ee M9 Se aGonclusionsty Acts tater. os. been f oeswe tie. 4 ssenuest Lene edey...:: 120 525 Overali@onclusion Limitationsand PuriresResearchuwies. am, 0. 6.........- al

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Appendix Attam pnd Siete! Deu wires eo Vetinbics fom. TPR aed, Fs .. 127

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Table 1.1 Table 3.1

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Table 4.5

List of Tables

Dime tureiandsName ofthat ywAciisinn .ona.L0.D: Sample, Per. S.aclvect.. Average Daily Nutrient Intake for Children Aged 4-7 Assessed by 3-day Hoocthecord same een ae soanneres Atk LLLP Soepevherrt Gah winenecwevienan Average Daily Vitamin and Mineral Intake for Children aged 4-7

PAS CESSC CADE SECA VER OOCURCCOLUS rg trhc eta r ashe. ait ene emia C aN ae Average Daily Fatty Acid Intake for Children aged 4-7 Assessed by 3- GAVer COC SR CCOLCS) seamen mente Sanat lnts i aa ee ene ee meiene Wgn Weight, Height, and Body Mass Index of Subjects in Control and LCP Groupsatqand.y sVlOntinsaeeene ess f75ys ace ohana k ee eee seen, So Daily Nutrient, Vitamin and Mineral Intake from Food Consumption Assessed using 4-day Food Record at 0 and 7 months of Subjects in Controlancd: LE GPr Groups mee esen wun cc oeees oats ae pa ee ee Daily Fatty Acid Intake of Food Consumption Assessed Using 4-day Food Record at 0 and 7 Months of Subjects in Control and LCP

Comparison of Docosahexaenoic Acid Intake Determined From 4-day Food Record and Food Frequency Questionnaire at 0 and 7 months in @ontroland: | @ESSublecise sean ca at states a cote. sone eee eeneen a: Mean and Standard Deviation of Variables from TVPS-R at 0 and 7 Month Comparisons of Subjects in Control and LCP Groups...............

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Table B.1 Nutrient Composition of Control and LCP Supplement fed.............

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Besentia PattveAcrdbvietabolisiisess. Visti Aacite it elites... Distribution of AA Intake (mg/day) Assessed by 3-day Food Records fom@hiidrentnge d "4 Siemens... ate aes eis Cour... Distribution of DHA Intake (mg/day) Assessed by 3-day Food Records Loun@ in ldneneA Ged <peet ie Sas RN ITN Suchen vo: csccrtsn vite dscclous ableloaatidas Mean Individual Intakes of AA (mg/day) from Food Consumption Assessed by 4-day Food Records at 0 and 7 Months in Control and LCP SULT OC steers emer rare mnt te Sede c aceefuiay'niy Hy coef au eats a Mee ore RR Mean Individual Intakes of DHA (mg/day) from Food Consumption Assessed by 4-day Food Records at 0 and 7 Months in Control and LCP SOUT CCLS Seem ere to ee ere a Ras Fee BP Get ce Sc ae Wick ou ec een Re oING DHA Intake (mg/day) plotted versus Median Visual Perceptual age (ITORLS)) Oe eet e foc perenne Oc Hear ten eek heyy Okc ek Ghote aiah fae AA and DHA level in Plasma Phospholipids in Control and LCP SUDICCIS al wieIVIONUNS aceon cpt Ot ase A) oh met ene aener Wel et Da aeutn ee: AA and DHA level in Phospholipids of Red Blood Cells in Control and IGEASUDI eCiseate/ «NVLONIISMMeNee er ar. ota ine eatery ere ete en eee ones

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Appendix Figures

Figure D.1 Sample HOVT Chart used to Assess Visual Acuity in Children Figure D.2 Sample Lang Test used to Assess Stereo Acuity in Children............... Figure D.3 Sample Ishihara Pseudoisochromatic Plate used to Assess Colour NU CGT AD EALCH ATION EIAY. «oo, c,d esate ep are na ate SPE Rte dae aan apa cn re ae Mrere:-45 Prism Cover Test used tovAssessiStrabismustratt 1. elas, Ae Figure E.1 Sample Graphical Representation of Each of the 7 Subsets in the Test of Visual Perceptual Skills (non-motor)- Revised (TVPS-R)

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List of Abbreviations

Arachidonic Acid

Adequate Intakes

Acceptable Macronutrient Distribution Ranees Linolenic Acid

Analysis of Variance

Body Mass Index

Docosahexaenoic Acid

Estimated Energy Requirements Essential Fatty Acids

Eicosapentaenoic Acid

Linoleic Acid

Low Birth Weight

Long Chain Polyunsaturated Fatty Acids Polyunsaturated Fatty Acids

Red Blood Cell

Test of Visual Perceptual Skills (non-motor)- Revised

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Acknowledgements

I would like to express my sincere thanks to my supervisor Dr. M.T. Clandinin, for providing me with the opportunity to work on this project. Thank you for your guidance

and continuous support throughout this degree

Thank you to my committee members, Dr. CJ Field and Dr. IM MacDonald; they were

integral to the completion of my degree.

Thank you Mich Wilke for always listening to me and providing me with great advice, and thanks to Nancy Evans who was there to assist me during both research projects. I would also like to thank Dr. Goh for his technical assistance and statistical help. Thank you for your patience and always taking the time to answer all my questions. Thank you to the all the members of the Dr. Clandinin lab group for all their help, and thank you to the support staff of the Department of Agricultural, Food and Nutritional Science. I would also like to thank Dr. JR Michelson and Marni Pearce for helping out with the neuropsychological assessments, and thanks to Brad Wakeman and Michelle Lang who

performed all the visual function assessments.

I would like express thanks to my parents, Herbert and Diana Lien for their love and support. Thank you to my best friend Iva Seto; she was there with me from the start to finish and for the good times and the bad. Thanks to both my family and friends for

believing in me and getting me through the tough times.

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I would also like to thank the Superintendent, Principals and Teachers of Parkland School County, and all the parents and children in the research study. Without them, this

research would not have been possible.

This work was financially supported by Natural Sciences and Engineering Research Council of Canada, MTI Meta Tech Inc, and Wyeth Nutitionals Incorporated. Vanessa Lien was a recipient of the Natural Science and Engineering Research Council (NSERC) Postgraduate Scholarship, the Alberta Heritage Foundation for Medical Research Health

Research Studentship, and the Dietitians of Canada Graduate Memorial Award.

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Chapter 1- General Introduction

1.1 Introduction

Long chain polyunsaturated fatty acids (LCP), particularly arachidonic acid (AA) and docosahexaenoic acid (DHA) are important in infant growth and development (Clandinin et al. 1981). These fatty acids are major components of the membranes of the brain and retina (Sastry 1985; Tinoco 1982). Supplementing infant formulas with both AA and DHA or DHA alone, has shown improvement in vision and brain development in both preterm (Birch et al. 1992; Clandinin et al. 2005) and term infants (Agostoni et al. 1995; Birch et al. 2002). Infant formulas in North America now contain AA and DHA (Health Canada 2002; U.S. Food and Drug Administration 2002). Many other countries such as Europe, Middle East, South America, Australia, Japan, Thailand, and some other Asian countries, already have LCP added to many of their preterm and term formulas (Fleith and Clandinin unpublished).

Although the rate of growth slows, the brain and eye continue to develop throughout childhood (Chugani 1998; Dobbing 1972; Oyster 1999). However, there is limited research investigating vision and brain development beyond 2 years of age. This literature review will investigate the role of AA and DHA in retinal and brain development, current dietary intake of these fatty acids in children, and provide an

understanding of the importance of LCP in childhood.

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1.2 Omega 6 and Omega 3 Fatty Acids

1.2.1 Nomenclature

There are two families of polyunsaturated fatty acids (PUFA): omega 6 and omega 3 fatty acids. Fatty acids are named using four common systems: two abbreviations (n-designation, A-designation), systematic name, and trivial name. For the n-designation (also known as the omega (@) end of the chain), the carbon numbering starts from the methyl end of the fatty acid. This reference system indicates the total number of carbons atoms in the chain, the number of double bonds, and the position or carbon atom number of the first double bond in the chain. For example with 18:2n6 or 18:26, there are 18 total carbon atoms in the chain, there are two double bonds, and the carbon atom number of the first double bond in the chain denoted from the methyl end is 6 (Table 1.1). In the A-designation, this system indicates the total number of carbon atoms, the total number of double bonds, and the superscripted numbers after the delta system indicate the position of the carbon atoms numbered from the carboxyl end at which the double bond begins. For example, 18:2A” ac denotes that there are 18 carbon atoms in total, there are two double bonds, at the 9 and 12 position, counting from the carboxyl end. Another type of system is the systematic name, which follows the nomenclature from the International Union of Pure and Applied Chemistry (IUPAC-IUB Commission on Biochemical Nomenclature 1974). This system modifies the name of the straight chain hydrocarbon and has the same number of carbon atoms. Using the hydrocarbon name, it removes the final —e, and replaces it with a —oic followed by the word acid. The trivial (common name) is typically derived from the common source of

the compound or the source from which it was first isolated and gives no clues to the

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1.3 Essential Fatty Acids and Metabolic Pathways

PUFA, particularly linoleic acid (18:2n-6, LA) and linolenic acid (18:3n3, ALA), are considered precursors of longer chain omega 6 and omega 3 fatty acids (Connor and Neuringer 1988). The ‘parent’? PUFA are known as essential fatty acids (EFA) and are necessary in the diet as they cannot be synthesized in humans. Burr and Burr (1929; 1930) were one of the first to describe that fats were essential to the diet through the investigation of a fat-free diet in rats. When fat is excluded in the diet, a deficiency disease develops which in rats results in an early death, unless they are given a curative dose of fat (Burr and Burr 1929; 1930).

Symptoms of EFA deficiency include growth retardation (Caldwell et al. 1972; Evans et al. 1934a, b), impaired reproduction (Evans et al. 1934a, b), kidney damage (Holman 1968), skin (Hansen et al. 1958; T-W-Fiennes et al. 1973) and hair abnormalities (Caldwell et al. 1972; T-W-Fiennes et al. 1973), impaired immune function (Caldwell et al. 1972), premature death (Evans et al. 1934a, b), fatty liver (Holman 1968), degenerative changes in the lung, (Uauy et al. 1989), and increased metabolic rate (Uauy

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