Controlled Release of Antioxidants via Biodegradable Polymer Films into Milk and Dry Milk Products

173 Pages·2003·4.02 MB·English

Controlled Release of Antioxidants via Biodegradable Polymer Films
into Milk and Dry Milk Products

Marleen van Aardt

Dissertation submitted to the Faculty of the
Virginia Polytechnic Institute and State University
in partial fulfillment of the requirements for the degree of

Doctor of Philosophy
in
Food Science and Technology

Susan E. Duncan, Chair
Timothy E. Long
Joseph E. Marcy
Sean F. O’Keefe
Susan R. Nielsen-Sims

November 21, 2003
Blacksburg, Virginia

Keywords: Lipid Oxidation, Milk, Poly(lactide-co-glycolide), Antioxidants, a-Tocopherol,
Butylated hydroxytoluene, Butylated hydroxyanisole

Copyright 2003, Marleen van Aardt
Controlled Release of Antioxidants via Biodegradable Polymer Films
into Milk and Dry Milk Products

Marleen van Aardt

(ABSTRACT)

Thermogravimetric analysis (TGA) was used to determine the oxidative stability of edible oils
(olive oil, milkfat) and triacylglycerides (triolein, trilinolein), while the effect of natural (a -
tocopherol, ascorbic acid) and synthetic antioxidants (butylated hydroxyanisole (BHA),
butylated hydroxytoluene (BHT), and tertiary butyl hydroquinone (TBHQ)) were evaluated on
stability of trilinolein. Trilinolein was oxidatively less stable than triolein, olive oil, and milkfat,
while triolein was elss stable than olive oil and milkfat. When comparing effectiveness of
antioxidants, a combination of 0.01% BHA and 0.01% BHT increased trilinolein stability the
most.

Sensory testing showed that the addition of a combination of 0.025% a -tocopherol and 0.025%
ascorbic acid to milk did not influence milk flavor. However, when adding a -tocopherol and
ascorbic acid to milk at these levels, light-induced oxidation off-flavor was significantly reduced
in comparison to unspiked milk after 10 h of light exposure (1100-1300 lx).

The effect of antioxidants (added in a single initial dose, and in weekly additions) on light-
exposed extended shelf-life milk over six weeks of light-exposed storage was measured by
volatile analysis. An initial single addition of a -tocopherol and ascorbyl palmitate significantly
reduced hexanal content in light exposed milk throughout the first four weeks of storage,
whereas the weekly addition of a combination of BHA and BHT reduced heptanal content of
milk to such a degree that no significant difference was observed when compared with light-
protected control milk.

Poly(lactide-co-glycolide) (50:50) films loaded with 2% a -tocopherol and a combination of 1%
BHT and 1% BHA were used in an antioxidant release study in water and Miglyol 812® at 4oC
and 25oC, as well as a study on the effect of these films on dry whole milk and dry buttermilk
stability. BHT was released through the hydrolytic degradation of the polymer when stored in
water at room temperature for eight weeks. However, it was expected that hydrolytic polymer
degradation would not take place when antioxidant-loaded films were stored in dry whole milk
(3.01% moisture) and dry buttermilk (4.60% moisture). a -Tocopherol, BHA, and BHT migrated
into whole milk powder through diffusion over four weeks of storage at 25oC, and significantly
reduced light-induced oxidation compounds.
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ACKNOWLEDGEMENTS

In 1998 I arrived on the doorstep of the Food Science and Technology Department of Virginia
Tech with nothing more than my undergraduate degree and related experience from South
Africa. I was immediately welcomed and given the opportunity to further my academic and
professional development with the pursuit of a graduate degree. This opportunity is something
that I will never forget, and I owe special thanks to Dr. Cameron Hackney (then department
head) and my advisor, Dr. Susan Duncan. Dr. Duncan has been a true mentor, sometimes
leading, sometimes showing me the way, at other times giving me the freedom to explore, but
always available with wisdom, advice, and support. I am greatly indebted to her sustained effort
and guidance in helping me come this far.

I would also like to extend my deepest appreciation to the other members of my graduate
committee, Drs. Joe Marcy, Tim Long, Sean O’Keefe (all from Virginia Tech), and Susan Sims
(Eastman Chemical Co.). Each of these individuals contributed to my graduate career through
their support and expertise in their respective fields of knowledge. Their doors were always
open and without their input I would have been stuck at many dead ends.

Thank you very much also to the faculty of the Food Science and Technology Department of
Virginia Tech for their support and commitment to graduate excellence at Virginia Tech. I
would especially like to thank Dr. Susan Sumner (department head) for being approachable and
supportive of graduate students in the department.

To all my peers, students in this and other departments, I thank you from the bottom of my heart
for long days spent in discussion, helping each other, and the camaraderie we shared. I would
like to thank Sonia Gonzalez, Jenny Norton, Stephanie Penn, Maureen Akins, Janet Webster, and
Heather Clarkson for your wonderful support and the time spent in- and outside of the office.
May we always keep in contact and extend friendships forged here to span even across the globe!
My sincere thanks also go to the graduate students in the Chemistry Department, specifically
Jeremy Lizotte, Dave Williamson, and Ann Fornof for their invaluable help with chemical
analysis.
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Two people that were absolutely integral to my studies, Kim Waterman and Walter Hartman,
deserve special mention. Project support is the backbone of our efforts as graduate students;
these two individuals helped to keep lights burning, milk processed, results analyzed - they were
always there to keep the cogs moving. Thank you, Kim and Walter, for all your hard work and
support, I will never forget that.

To Wolfgang and Heidi Glasser, our family’s “foster parents” in Blacksburg, thank you very
much for evenings filled with long discussions on academics and life, for your friendship and
support, for making us feel welcome in Blacksburg, and for taking us under your wings. You
will always be remembered and thought of very fondly.

I would also like to thank my husband and best friend, Jan van Aardt, for his constant love and
support – for many interesting discussions about polymer degradation and “where did the
antioxidants go”, as well as caring extensively for our eight-month old, Karla. He helped my
dreams come true.

Warm thanks go also to my loving parents, Jonatan and Marietjie van Eeden, for their love and
support throughout the past five years in a foreign country.

My sincere thanks go to Dairy Management Inc. for financial support of this project.

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TABLE OF CONTENTS

Page

Title Page……………………………………………………………………………......................i
Abstract……………………………………………………………………………………………ii
Acknowledgements…………………………………………………………………………….....iv
Table of Contents………………………………………………………………………………....vi
List of Figures………………………………………………………………………………….…xi
List of Tables…………………………………………………………………………………....xiv

CHAPTER I
INTRODUCTION.........................................................................................................................1

CHAPTER II
LITERATURE REVIEW……………………………………………………………………….4
2.1 Lipid Oxidation……………………………………………..……………………..4
2.1.1 Initiation Reactions…………………………………..……………………5
2.1.2 Propagation reactions…………………………………..………………….5
2.1.3 Termination Reactions…………………………………...………………..6
2.2 Protein Oxidation…………………………………………………...……………..7
2.3 Photo-oxidation of Milk Components……………………………….……………8
2.4 Packaging Materials as Photo-Protectors……………………………...…………11
2.5 Measurement of Oxidative Stability……………………………………..………12
2.5.1 Peroxide and Hydroperoxide Analysis……………………………..……12
2.5.2 Carbonyl Analysis……………...……………………………………...…13
2.5.3 Oxygen Absorption Methods………………………………………….…13
2.5.4 Chromatographic Methods…………………………………………….…14
2.6 Food Antioxidants……………………………………………………………..…15
2.6.1 Primary Antioxidants………………………………………………….…15
2.6.2 Synergistic Antioxidants…………………………………………………16
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2.6.3 Antioxidants in Milk……………………………………………………..17
2.6.4 Legislative Aspects Concerning Food Antioxidants……………………..19
2.7 Poly(lactide-co-glycolide) in the Food Industry…………………………………20
2.7.1 Synthesis………………………………………………………...……….22
2.7.2 Polymer Characteristics…………………………………………...……..23
2.7.3 Degradation…………………………………………………………..…..24
2.7.4 Controlled Release…………………………………………………….....27
2.7.5 Diffusion Parameters………………………………………………….....28
2.7.6 Polymer Safety………………………………………………………..….29
2.8 References………………………………………………………………………..30

CHAPTER III
EFFECT OF ANTIOXIDANTS ON OXIDATIVE STABILITY OF EDIBLE FATS
AND OILS: THERMOGRAVIMETRIC ANALYSIS……………………………………….36
Abstract………………………………………………………………………………..…37
Introduction………………………………………………………………………………38
Materials and Methods………………………………………………………………...…40
Materials and Sample Preparations………………………………………………40
Fatty acid Profiles…………………………………………………………..……40
Thermogravimetric Analysis………………………………………………...…..41
Statistical Analysis…………………………………………………………….....42
Results and Discussion…………………………………………………………………..43
Conclusion ………………………………………………………………………………48
Acknowledgements………………………………………………………………………48
References………….…………………………………………………………………….49

CHAPTER IV
LIGHT-INDUCED FLAVOR OF MILK FORTIFIED WITH a-TOCOPHEROL
AND ASCORBIC ACID............................................... ……………………………………….57
Abstract………………………………………………………………………………..…58
Introduction………………………………………………………………………………59
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Materials and Methods………………………………………………………………...…62
Milk Processing…………………………………………………………...……..62
Preparation of Antioxidant-Spiked Samples……………………………………..62
Milk Storage and Handling………………………………………………...….....63
Sensory Analysis…………………………………………………………………63
Triangle test for Similarity…………………………………………...…..64
Triangle test for Difference………………………………………………64
Chemical Analysis……………………………………………………………….65
Gas Chromatography-Olfactometry..……………………………………65
Training of Panelists for GC-O………………………………………….66
Thiobarbituric Acid Reactive Substances Test (TBARS)……………….66
Statistical Analysis…………………………………………………………….....66
Results and Discussion…………………………………………………………………..68
Sensory Analysis………………………………………………………………....68
Chemical Analysis……………………………………………………………….70
Conclusion ………………………………………………………………………………73
Acknowledgements………………………………………………………………………73
References.……………………………………………………………………………….74

CHAPTER V
AROMA ANALYSIS OF LIGHT-EXPOSED MILK STORED WITH AND WITHOUT
NATURAL AND SYNTHETIC ANTIOXIDANTS........…………………………………….81
Abstract………………………………………………………………………………..…82
Introduction………………………………………………………………………………83
Materials and Methods………………………………………………………………...…86
Milk Processing…………………………………………………………...……..86
Preparation of Antioxidant-Spiked Milk Samples…….…………………………86
Storage and Light Exposure……………………………………………...…........87
Microbiological Analysis………………………………………………………...87
Volatile Analysis…………………………………………………………………88
Lipid Extraction and HPLC Analysis of Antioxidants…………………………..88
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Statistical Analysis…………………………………………………………….....89
Results and Discussion…………………………………………………………………..90
Microbiological Analysis………………………………………………………...90
Extent of Lipid Oxidation: Volatile Analysis…………………………................90
Lipid Oxidation Odors…………………………………………………………...92
Antioxidant Content in Control- and Antioxidant-Treated Milk………………...94
Conclusion ………………………………………………………………………………96
Acknowledgements………………………………………………………………………96
References………….………………………………………………………………….…97

CHAPTER VI
ANTIOXIDANT-LOADED BIODEGRADABLE FILMS: CONTROLLED RELEASE
OF ANTIOXIDANTS INTO DRY MILK PRODUCTS AND FOOD SIMULATING
LIQUIDS........……………………………………………………………………………….…109
Abstract…………………………………………………………………………………110
Introduction……………………………………………………………………………..112
Materials and Methods………………………………………………………………….115
Materials…………………………………………………………...…………...115
Antioxidant-Loaded PLGA films…….………………………………………...115
Characterization of Polymer Films…………………………………...…...........116
Characterization of Dry Whole Milk and Dry Buttermilk……………………...116
Release of Antioxidants and Degradation of PLGA films in Water and
Miglyol 812®……………………………………………….…………………...117
Volatile Analysis………….…………………………………………………….117
Lipid Extraction and HPLC Analysis of Antioxidants in Powders…………….118
Statistical Analysis …..……………………………………................................118
Results and Discussion…………………………………………………………………120
Release of Antioxidants and Degradation of PLGA Films in Water and
Miglyol 812®……………………………………………………………………120
Release of Antioxidants and Degradation of PLGA Films in Dry Whole Milk
and Dry Buttermilk……………………………………………………………..123
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Conclusion ……………………………………………………………………………..126
Acknowledgements……………………………………………………………………..126
References………………………………………………………………………............127

APPENDICES…………………………………………………………………………………140

Appendix A: Human Subjects Forms for Sensory Evaluation…………………………………140
Appendix B: Sensory Test Scorecard…………………………………………………………..143
Appendix C: Worksheet for Sensory Triangle Test for Similarity……………………………..144
Appendix D: Bacterial Counts (CFU.ml-1) for Extended Shelf-life (ESL) Milk Treated
With Single- and Weekly Additions of Antioxidants………………….…………...…..145
Appendix E: Odor-Active Compounds Detected by GC-O of Control and Antioxidant-
Treated Milk when Exposed to Light (1100-1300 lx) for 12 h per day for 6 weeks…...146
Appendix F: Antioxidant Content of Antioxidant-Treated Milk During 6 Weeks of Storage…152

CURRICULUM VITA………………………………………………………………………..154
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