IEEE Life Sciences Grand Challenges Conference December 2-3, Singapore U-Town, National University of Singapore

THE WANDERINGS OF A FREE

Professor Barry Halliwell Tan Chin Tuan Centennial Professor Deputy President (Research & Technology) National University of Singapore Life history

Year Event

1968 - 1971 Studied Chemistry and Biochemistry at St Catherine’s College, Oxford University 1971 - 1973 Stayed on to do a D.Phil on plant metabolism, Botany School. Studied photorespiration. (a rather well worn topic)

1974 - 1988 Lecturer and subsequently Reader, Biochemistry, Kings College London.

1988 - 2000 Professor, Division of Pharmacology, Kings College London 1995 – 1998 Decided to try something new (one year full time, almost Visiting Research Professor in Internal Medicine and Biochemistry, four years part time) University of California, Davis / Berkeley 1998 - 2000 Decided to try something new again Visiting Professor of Biochemistry, NUS 2000 - 2007 Head, Department of Biochemistry, NUS

2003 - 2008 Executive Director, NUS Graduate School for Integrative Sciences & Engineering 2006 - present Deputy President (Research & Technology)

WHAT NEXT?

Plants are essential to humans • Supply oxygen • Supply diet-derived (ascorbate*, tocopherols*, carotenoids,? Flavonoids?) • Diets rich in plants are associated with lower disease risk (some cancers, diabetes, cardiovascular, dementia) But why?

THE LUCK OF THE DRAW

BETTER THAN PHOTORESPIRATION ALL ASPECTS OF AEROBIC LIFE INVOLVE FREE RADICALS AND ANTIOXIDANTS – YOU CANNOT ESCAPE THEM, NOR SHOULD YOU WISH TO.

Halliwell B. (2006) Reactive species and antioxidants. biology is a fundamental theme of aerobic life. Plant Physiol. 141, 312-22

Oxidant – Balance Peroxiredoxins AA Metabolism system

Phagocytes SOD, Catalase

Diet-Derived Antioxidants

Mitochondrial Blood respiration components NADPH oxidases Iron chelators Albumin, DUOxes Caeruloplasmin, Xanthine oxidase Transferrin, haptoglobin etc So some oxidative damage is inevitable and repair &/or replacement of damaged molecules is essential ROS are bad They contribute to many age-related diseases and possibly to the ageing process itself. But they are also good, they help protect against disease in animals and coordinate the inflammatory response, e.g. via redox- signalling, as well as participating in the killing of pathogens. Phagocyte ROS production is a key event.

NEURODEGENERATIVE DISEASES ARE ACCOMPANIED BY

Amyloid plaques Neurofibrillary tangles 1. Mitochondrial Alzheimer disease Alzheimer disease dysfunction 2. Accumulation of abnormal proteins Lewy Body Inclusion bodies 3. Increased oxidative Parkinson disease Huntington disease damage 4. Iron deposition OXIDATIVE DAMAGE IN ALZHEIMER DISEASE Rises in 8OHdG and other base oxidation products in both nuclear and mitochondrial DNA, also rises in 8OHG, protein carbonyls, glutamic and aminoadipic semialdehydes (protein oxidation products), methionine sulphoxide, nitrotyrosine, acrolein, HNE, HO-1 and IPs in brain tissue. Plaques and paired helical filaments contain oxidized, glycated, and nitrated proteins, and CSF from AD patients is also reported to contain elevated levels of oxidized, nitrated and glycated proteins. One oxidized protein associated with plaques is CuZnSOD. Damage is already elevated in brain tissue and CSF in mild cognitive impairment and early AD.

Current Research in the Halliwell Lab 1. Antioxidants in Human Health and Disease 2. Mechanisms of Neurodegeneration 3. Artefacts in Cell Culture 4. Ageing in the nematode C.elegans WHY RESEARCH IN SINGAPORE?

South Korea Japan Middle East China

India Bangladesh Hong Kong Taiwan

Thailand Vietnam USA Europe Sri Lanka Philippines Malaysia Australia Singapore Brunei Advantages • No energy (except some solar) • Location • Little food New Zealand • Political / social stability • Little space • Stable government • No oil or mineral resources Indonesia • People / High level of • Water-constrained education • Climate change • Connectivity • Very small, minute domestic market • Good social coherence • Low reproduction rate • Globally connected means risk of disease / financial crisis etc can impact very fast STRENGTHS OF NUS

Research is conducted in All Major Disciplines Faculties and Schools (Undergraduate and Graduate education) 1. Arts and Social Sciences 7. Law 2. Business 8. Medicine 3. Computing 9. Music 4. Dentistry 10. Public Health 5. Design and Environment 11. Science 6. Engineering 12. University Scholars Programme (for Undergraduate only)

Graduate Schools 1. Duke-NUS Graduate Medical School 2. Lee Kuan Yew School of Public Policy 3. NUS Graduate School for Integrative Sciences and Engineering RESEARCH INSTITUTES & CENTRES 23 University-level RICs + 3.5 Research Centres of Excellence

• Asia Research Institute • Mechanobiology Institute • Cancer Science Institute of • Middle East Institute Singapore • NUS Environmental Research Institute • Centre for International Law • NUS Nanoscience & Nanotechnology • Centre for Maritime Studies Initiative • Centre for Remote Imaging, Sensing • NUS Research Institute in Suzhou & Processing • Risk Management Institute • Centre for Quantum Technologies • Singapore Centre on Environmental • East Asian Institute and Life Sciences Engineering • Energy Studies Institute • Singapore Institute for Neurotechnology • NUS Global Asia Institute • Singapore Synchrotron Light Source • Institute for Mathematical Sciences • Solar Energy Research Institute of • Institute of Real Estate Studies Singapore • Institute of South Asian Studies • Temasek Laboratories • Interactive & Digital Media Institute • The Logistics Institute-Asia Pacific • Life Sciences Institute • Tropical Marine Science Institute

*Occupants SINGAPORE’S NATIONAL AND ONLY MIT COMPREHENSIVE UNIVERSITY ETH Zurich TUM Munich Synergy in Proximity Imperial College Hebrew University Technion Israel Berkeley Peking University CREATE Biopolis NUS NTU Others Fusionopolis

National University SICS Hospital MRT! one-north

Map courtesy of Singapore JTC Corporation Science Parks

15 SICS – Singapore Institute for Clinical Science (A*STAR) Duties of DP(R&T) Office - Administration and Compliance Promote NUS-Industry Exchange - Facilitation Grant Seed Grant Dialogue with Funding Administration Matching Grant Scheme Funders Identify Areas of Strategic Importance High Impact Growing Research the Pie Promote Multidisciplinary Research Programmes Attract & Retain Talent  Build solid base of high-quality research across a reasonably-broad range of disciplines Research Strong Global Benchmarking  Establish Research Centres of Excellence Profile Review within NUS & other Peaks of International Research and relative to peer Excellence in selected areas universities (research benchmarking) Recognition & Establish International Facilitate Commercialisation Research Networks of Research Outcomes Research Reward of Research Spin Off Excellence Publicise Achievements

IP Protection Protection of Research Prestigious Research Awards Integrity Animal Welfare Institutional Review Board (IACUC) NUS CAN NURTURE FOR SINGAPORE NICHE AREAS OF HIGH QUALITY THAT ARE NOT YET THE “FLAVOUR OF THE MONTH” [e.g. non-medical biology, plant science, humanities and social science (e.g. Asia Research Institute), pure mathematics] One example • Molecular basis of crop yields (MOU signed with the International Rice Research Institute (IRRI) on 16 Feb 2009) • Crop resistance to environmental change • Nutrition, diet and health maintenance in Asians • Biodiversity

• New competitive grants from NRF and SMF (3 grants totalling $21.2 million were obtained in the food security area)

An example of how “basic” research suddenly becomes relevant!

BUT SINGAPORE IS VERY PRAGMATIC Demonstrate your value

WHAT IS NUS FOR?

• Universities in Singapore are complementary to A*STAR research institutes.

• One key function of research intensive universities is to provide a strong and broad research base not only to address concerns of today but to create opportunities for tomorrow.

• A university PI recruited on the basis of his/her promising and novel research concepts typically begins as Assistant Prof to build his/her research and compete for small grants (<$1M). Over time and with more experience, he/she will develop a unique research niche and also will gain the strength to participate in bigger programmes and consortia. NUS SEEKS TO CONDUCT IMPACTFUL RESEARCH WHAT IS IMPACT OF RESEARCH? • Outstanding fundamental research of high intellectual impact that attracts attention to Singapore as a country capable of performing such research and grows NUS’ global reputation • Research which helps to grow new industries for Singapore and to develop existing ones, e.g. by spin-offs and licensing of Intellectual Property (IP) • Research that helps to attract high-level foreign industry to locate in Singapore • Research that makes Singapore a better place to live and improves the health and welfare of the population • Research that expands intellectual breadth and develops ideas and discourses about human experiences which will prepare us more effectively for an increasingly global and cosmopolitan world • Research that influences and informs government policy • Research that enhances the security of Singapore (e.g. defence, food, energy supply) Note that the best research programmes often contribute in several ways.

When I was young, the Free Radical –Antioxidant Field was SIMPLE

 Free radicals are bad  Antioxidants are good  Taking antioxidants will prevent disease  Since free radicals are implicated in ageing, antioxidants will make you live longer

When I was young, it was SIMPLE Taking Antioxidants will Prevent Disease

(with several exceptions) Intervention Trials with high-dose Vitamin E Limited efficacy in Alzheimers disease No effect in mild cognitive impairment No effect on cardiovascular disease No beneficial effect on cancer Similar failures with and β-carotene Suggestions of deleterious effects in some studies, e.g. increased risk

BMJ (2010) 341:c5702 Conclusion In this meta-analysis, vitamin E increased the risk for haemorrhagic stroke by 22% and reduced the risk of ischaemic stroke by 10%. This differential risk pattern is obscured when looking at total stroke. Given the relatively small risk reduction of ischaemic stroke and the generally more severe outcome of haemorrhagic stroke, indiscriminate widespread use of vitamin E should be cautioned against. Intervention Trials with Vitamin E

• Vitamin E works much better in (at least some) rodent models of neurodegeneration (e.g. APP, ALS) and cardiovascular disease • And it also decreases oxidative

damage (measured as F2-isoprostanes) in these models

VALIDITY OF RODENT MODELS OF STROKE AND NEURODEGENERATIVE DISEASE?

By courtesy of John Milner, National Cancer Institute THERE ARE MULTIPLE EXPLANATIONS FOR THIS “FAILURE” OF ANTIOXIDANTS LET’S RE-EXAMINE OUR STARTING HYPOTHESIS • Steady-state levels of oxidative DNA damage contribute significantly to the major cancers. (PROBABLY TRUE) Ref: Halliwell B (2007) Oxidative stress and cancer: have we moved forward? Biochem. J. (2007) 401, 1-11. • Steady-state levels of lipid peroxidation contribute significantly to cardiovascular disease (POSSIBLY TRUE) and to neurodegenerative disease (PROBABLY TRUE) Ref: Halliwell B (2006) Oxidative stress and neurodegeneration; where are we now? J. Neurochem. 97, 1634-58.

IN CARRYING OUT THE INTERVENTION TRIAL, DID WE DECREASE OXIDATIVE DAMAGE IN THE SUBJECTS?

Because if we did not, no effect would be predicted This was assumed, but not tested Our Studies and Many Others using Biomarkers of Lipid Peroxidation and Oxidative DNA Damage show NO DECREASE on Supplementing Healthy Volunteers with Ascorbate, Beta-Carotene or Vitamin E. If antioxidants rarely change oxidative damage levels in humans, what does? • Obesity (Humans) • Hyperglycaemia (Humans) • High plasma LDL Cholesterol (Humans) • High Cholesterol Diet (Rabbits and rats , humans probably not) • Zinc Intake (Rabbits, some other animals, human data mixed) • Body Iron Levels (Rabbits, rats, mice, maybe humans) • Certain foods (Humans, e.g. dark soy sauce, tomato) • Diabetes (in some studies, not others)*, but probably not the metabolic syndrome

• Intake of polyunsaturated fatty acids (PUFAs) ∆ (docosahexaenoic acid, possibly eicosapentaenoic acid)

• Cigarette smoking • Sunlight (eye, skin)

• Other environmental toxins?? (O3, NO2, arsenic in water) *May depend on how well glucose and lipids have been normalized in the diabetic cohorts studied ∆Despite the propensity of PUFAs to oxidise in vitro, growing evidence suggests that they minimise oxidative damage in vivo.

Halliwell B (2011) Free radicals and antioxidants: - quo vadis? Trends Pharmacol. Sci. 32:125-130.

STRATEGY TO MINIMISE OXIDATIVE DAMAGE SCENARIO SUGGESTED STRATEGY Obesity Lose weight Hyperglycaemia Control blood sugar High plasma LDL Cholesterol Control LDL cholesterol

Body Iron Levels Avoid iron supplements Certain foods (Humans, e.g. dark soy sauce, Eat diet rich in fruits and vegetables tomato) Intake of polyunsaturated fatty acids (PUFAs) Eat fish regularly ∆ (docosahexaenoic acid, possibly Don‘t smoke eicosapentaenoic acid) (humans) It is essential to do appropriate controls in testing effects of foods, because the consumption of any food (antioxidant or not) can sometimes alter levels of certain biomarkers. ∆ Despite the propensity of PUFAs to oxidise in vitro, growing evidence suggests that they minimise oxidative damage in vivo.

Halliwell B (2011) Free radicals and antioxidants: - quo vadis? Trends Pharmacol. Sci. 32:125-130. Halliwell B (2012) Free radicals and antioxidants: updating a personal view. Nutr. Rev. 70:257-265.

CONCEPT OF HORMESIS ROS – Antioxidant Balance Peroxiredoxins AA metabolism Upregulation by Dietary Glutathione system Phagocytes pro-oxidants! SOD, Catalase Diet-Derived Antioxidants Mitochondrial Blood respiration components Iron chelators Albumin, Caeruloplasmin, Xanthine oxidase transferrin DO POLYPHENOLS ACT AS ANTIOXIDANTS &/OR PRO-OXIDANTS IN THE GASTROINTESTINAL TRACT, AND DOES IT MATTER WHICH? Several dietary constituents (e.g. sulphoraphane) upregulate endogenous antioxidant defences. LIFE AND BIOMEDICAL SCIENCES CLUSTER NUS Life Sciences Institute (LSI) NUS Centre for Life Sciences (CeLS) Competitive Space for Integrated Life Science Programmes Research Programmes in Life Sciences Disease-related themes Underpinning Science & Technology

. Cancer . Molecular Epidemiology / Genetics . Neurodegenerative disease Bioinformatics / Tissue Respository

. Vascular Diseases . Bioengineering/ Neuroengineering / . Infectious Diseases Tissue Engineering

. Human nutrition / disease . Medicinal Chemistry / Toxicology / revention Clinical Trials

. Healthy ageing . Structural Biology . Environmental microbiology . Lipidomics . Immunology

. Neuroscience, . Psychology / human cognition neuroengineering and cognition ALL CROSS-FACULTY, CROSS-DISCIPLINARY What is our Translational Medicine niche? • Preferred site in Asia for validation & testing in man of new diagnostics, drugs & devices for diseases that are more common in Asia, or where symptoms, outcome and pathology are different from the rest of the world • Deep expertise in disease biology and world-class Proof-of-Concept & early phase clinical trial capability with international accreditation

• Close link of basic biomedical research, engineering,*. and computing with clinical medicine • Differences between different ethnic groups

NUS Centre for Translational Medicine • 41,000 sqm • 15 floors: 9 floors for Research, 6 floors for Education • Clinical Imaging Research Centre • Investigational Medicine Unit • Cancer Science Institute Singapore • Other major programmes, including cardiovascular medicine, neuro-cognition, eye research, clinical nutrition and metabolic medicine (diabetes and obesity) Lung Cancer in Asians may be different from the West

Slide courtesy of Prof John Wong Vorinostat Histone deacetylase inhibitor under evaluation as anti-cancer drug

Glucuronidation (UGT2B17) Vorinostat Glucuronide (less activity) Vorinostat 4-Anilino-4-oxobutanoic acid Hydrolysis followed by β-oxidation

UGT2B17*2 (del variant) •Reduced UGT2B17 activity •Homozygotes: ∼ 10% Caucasians ∼ 60-70% Asians

UGT2B17*2 homozygotes present in 62% of our cohort

Wong et al. Pharmacogenetics & Genomics Slide courtesy of Prof John Wong 2011, 21(11):760-8 Asian Studies at NUS

Faculty of Arts and Social LKY School of Public Policy Sciences: Aim Asia Competitiveness Institute Research Clusters Centre for Asia and Globalisation

Cities Institute of Water Policy Environment To develop Family, Children & Youth special Health expertise, East Asia Institute Migration insights and Research on aspects of China: Religion connections Political, economic & social changes Science , Technology & Society relating to key Regional & global implications of issues of critical economic rise of China importance to Cultural & commercial networks of Asia Research Institute Asia ethnic Chinese Migration & human mobility Urbanisation Institute of South Asian Changing family structure & roles Studies Research on aspects of South Asia: Cultural studies NUS – Global Asia Economics & trade policy Religion & globalisation Institute Multilateral & international linkages Science, Technology & Society Asian Cities in context of Asia Politics & governance Life cycle financing Security NIHA (NUS Initiative to South Asia & economic change Improve Health in Asia) System dynamics NUS ALSO STRONGLY SUPPORTS RESEARCH IN HUMANITIES / SOCIAL SCIENCES / LAW / BUSINESS

• Important in its own right, e.g. to develop understanding and explanations of human conditions and behaviour. • Contributes to cross-disciplinary initiatives (environment, sustainability, digital media, ethics, risk management, ageing etc) • Holistic education of students Research at NUS addresses Singapore Problems LOOKING FORWARD - be ahead of the pack Challenges Facing Singapore

Energy (more efficient usage, securing supply)

Environmental management / global warming

Risk of infectious diseases

Securing the food supply / human nutrition

Ageing and age-related disease

World insecurity / financial risks in Asia

Sustainable cities Interdisciplinary Research, the Finance Cluster • NUS Global Asia Institute • Risk Management Institute • Institute of Real Estate Studies • Centre for Quantitative Finance • Centre for Asset Management Research & Investments • Centre for Behavioral Economics • Singapore Centre for Applied and Policy Economics • Office of Risk Management Prof HO Teck Hua Vice-President (Research Strategy)

Professor Ho Teck Hua is in charge of overseeing and building the University's Finance and Risk Management integrative research cluster. He concurrently holds the Tan Chin Tuan Centennial Professorship. Prof Ho has been a consulting professor to the NUS Overseas College in Silicon Valley since 2002.

He received a B.S. with first-class honours in Electrical Engineering (1985) as well as an M.S. in Computer and Information Sciences (1989) from the National University of Singapore. Additionally, from The Wharton School, University of Pennsylvania, he received an M.A. (1991) and a Ph.D. (1993) in Decision Sciences.

Prof Ho is currently the William Halford Jr. Family Professor of Marketing, and the Chair of the Marketing Department at the Haas School of Business at the University of California, Berkeley. Ho has been a chaired professor at U.C. Berkeley's Haas School of Business from 2002, and is also the Director of the Asia Business Center at the Haas School of Business from October 2007. Ho earned his tenure at The Wharton School, University of Pennsylvania in 1999. He was Assistant Professor of Operations and Technology Management at the UCLA Anderson School of Management from 1994-1997.

NUS ALSO STRONGLY SUPPORTS RESEARCH IN HUMANITIES / SOCIAL SCIENCES / LAW / BUSINESS

• Important in its own right, e.g. to develop understanding and explanations of human conditions and behaviour. • Contributes to cross-disciplinary initiatives (environment, sustainability, digital media, ethics, risk management, ageing etc) • Holistic education of students NUS-KEPPEL CORPORATE LAB

Partnership between NUS and KOMTech (Keppel Offshore Marine Technology Centre – R&D arm of Keppel OM). Funded by NRF Corporate Lab scheme. NUS-Keppel Lab will involve 30 staff from NUS (including 4 Visiting Profs) from CORE, FOE (CEE, ME, ECE, BME), TMSI and DBS. Coordinated by MI@NUS. KOMTech will have 19 staff involved. Total no. of researchers planned is 57. Research at NUS-Keppel Lab will focus on developing: • Innovative systems for deep and ultra-deep water and the Arctic for oil and gas exploration • More efficient and advanced production methods to replace or enhance manual operations in shipyards • Core competencies in environmental impact assessment and environmentally benign exploration and technology to harvest polymetallic nodules from seabeds at depths of 4000 to 5000m.

A COMING PROBLEM FOR SINGAPORE

Proportion of population aged 65+ in selected IARU countries

UK

DK

AUST 2030 2005 SG 1980

JP

CH

0 5 10 15 20 25 30 35

REUTERS/CORBIS Slide by courtesy of Dr Kenneth Howse, Oxford Centenarians now constitute the University fastest-growing age group owing to Source: UN Population database advances in health care. The International Alliance of Research Universities (IARU) is a collaboration between ten of the world’s Source – Nature 467 (2010), 274- leading research-intensive universities who share 275 similar visions for higher education, in particular the education of future leaders. IARU comprises ANU, ETH Zurich, NUS, Peking, Berkeley, Cambridge, University of Copenhagen, Oxford, University of Tokyo and Yale University. (Virtual) Institute for the Study of Ageing (VISA) . Anti-aging medicine (ethical) . Basic aging / Neurobiology research . Health care delivery / outcomes . Ageing & Lifestyle (nutrition, exercise etc) . Social aspects (e.g. community support) . Housing for the aged . Public policy (e.g. pensions) . Products for the aged . Dementia centre . City design (e.g. public transport) . Gerontology group NUS Schools and Faculties / Research Institutes/ Centres Dementia Centre Singapore Institute Human studies for Clinical Subject cohorts Sciences VISA Mild cognitive impairment Human studies Lifestyle and disease (NUHS) Industry liaison prevention Translational medicine/ Optimal environment nutritional products (ageing in place) Thought leadership for Government and Social sciences Basic Science charities Humanities Disease-related Public policy research Tsao Foundation Cognitive assessment Duke-GMS (NUHS) Global Asia LKY SPP Institute Financial / risk Exploring the identity management of the 21st century Asia city Healthcare policies Financing the elderly What does VISA aim to do?

• Biological determinants of ageing well

• Environments that best support ageing well

• Fiscal, medical & other policy issues that can be optimised to better support Singaporean ageing population Free radical theory of ageing 1956 Harman  Ageing results from random deleterious damage to tissues by free radicals.  Later the key role of mitochondria was emphasized  Superficially attractive because it

• Explains age – relation of diseases such as cancer and cardiovascular (oxidative damage involved) • Explains inverse correlation with basal metabolic rate (lower ROS burden per gram of tissue in bigger animals since they

consume less O2 per gram of tissue). • Accounts for the random element in ageing C.elegans as a Model System

• Soil dwelling nematode worm • Eats bacteria • 99% Self-fertilizing hermaphrodites

• => Isogenetic clones !

Adult: length: 1mm Ø: 100µm Volume: approx 1nL

Metabolic rate: 45nW Offspring: ~300 in < 8 days

Lifespan: max 40 days

WORK WORM OF AGEING RESEARCH Some more advantages • Extensive set of mutants and tools - 1000s of mutant strains PC - GFP reporter gene system - RNAi is relatively easy • Very “simple” nervous systems • Total complement of 302 neurons – Essentially invariant structure – Complete wiring diagram Control 10μg/ml 100μg/ml known • There is still randomness in the ageing process.

C PC 10μg/ml

C- Lifespan and Healthspan

Histogram for day of death for 200 isogenic C. Motility phenotype distribution in the same elegans clones, born on the same day in our lab and isogenic C. elegans population on different grown under identical culture conditions. Even days of life. Even thought both genome and though their genome as well as environment are environment are identical, surviving worms identical, their individual lifespan ranges from 10 of the same age may be found to have days to 34 days, probably due to intrinsic stochastic dramatically different motility class, an factors inherent in ageing. indicator of health. Artefacts in Cell Culture Cell Culture is an Oxidative Stress • Trypsinization  GSH • Serum deprivation  ROS • The major factor – OXYGEN

Intracellular pO2 (except skin, respiratory tract, cornea) 1 – 10 mmHg

95% air / 5% CO2 is ~150mmHg Therefore more ROS are made! • Oxygen levels also fluctuate wildly during culture and cell handling and with depth of medium Cell Culture is an Oxidative Stress Second major factor – cell culture media are poor in antioxidants • vitamin C • vitamin E (little lipid) • Selenium

DOES VITAMIN C CURE CANCER?

Clement, MV, Ramalingam, J, Long, LH and Halliwell B. The in vitro cytotoxicity of ascorbate depends on the culture medium used to perform the assay and involves . Antiox. Redox Signaling, 3, 157-163 H2O2 Generation by Ascorbate Long, LH, Clement, MV, and Halliwell B. Artifacts in cell culture: Rapid generation of hydrogen peroxide on addition of (-) – epigallocatechin, (-) – epigallocatechin gallate, (+) – catechin and quercetin to commonly used cell culture media. Biochem. Biophys. Res. Commun. 273, 50-53

Long LH, Kirkland D, Whitwell J, Halliwell B. (2007) Different cytotoxic and clastogenic effects of epigallocatechin gallate in various cell culture media due to variable rates of its oxidation in the culture medium. Mutat Res/Genet. Toxicol. Environ. Mutagen. 634, 177-183 COMPOUNDS THAT COMPOUNDS THAT DO NOT GENERATE H2O2 GENERATE H2O2

Apigenin Curcumin Delphinidin chloride

Rosmarinic acid Hesperetin Resveratrol

Naringenin Tyrosol Hydroxytyrosol Rapid loss of polyphenols upon incubation in DMEM at 37oC. Data are mean ± SD, n ≥ 3.

Delphinidin chloride Resveratrol

Hydroxytyrosol Curcumin

Long LH, Hoi A, Halliwell B (2010) Instability of, and generation of hydrogen peroxide by, phenolic compounds in cell culture media (2010) Archives of Rosmarinic acid Biochemistry and Biophysics. 501:162-169 Examples of artefacts caused by oxidation of compounds added to cell culture media

Halliwell B. (2008) Are polyphenols antioxidants or pro-oxidants? What do we learn from cell culture and in vivo studies? Arch Biochem Biophys. 476, 107-112 Thank You Questions & Answers

VISION MISSION

A leading global university To transform the way people centred in Asia, influencing think and do things through the future education, research and service

58