ANTIOXIDANT

Antioxidants The chemical compounds which can delay the start or slow the rate of lipid oxidation reaction in food systems.

Mechanism of Antioxidant 14 13 12 11 10 9 C H ( C H ) C H C H C H C H C H C H C H R 3 2 3 2 2 2 Initiation Metal Energy Reactive oxygen species Lipoxygenase Substrate effect -  H 13 12 11 10 9 C H ( C H ) C H C H C H C H C H C H R 3 2 4  2 E0= 600mv Oxygen consumption, Conjugated diene Electron spin resonance + 3O2 K=109/sec

. C H ( C H ) C H C H C H C H C H C H R Propagation O 13 12 11 10 9 C H ( C H ) C H C H C H C H C H C H R 3 2 4 2 Propagation O (K= 10o M-1sec-1) +  H from RH (triglyceride) O R.  E0=1000mv (K= 107 M-1sec-1) . O H O C ( C H ) +  H from 3 3 C ( C H ) 3 3 O C H 3 O C H 3 E0 = 300-500mv

Most reactive oxygen species 13 12 11 10 9 C H ( C H ) C H C H C H C H C H C H R 3 2 4 2 O Peroxide value O -  OH Most reactive oxygen species Transition Metal H E0=2300 mv 13 12 11 10 9 C H ( C H ) C H C H C H C H C H C H R 3 2 4 2 O  E0=1600 mv Termination Sensory evaluation Volatile compounds C H ( C H ) C HO 3 2 4 C H ( C H ) C H 3 3 2 3

Are you ready to fight the attack of prooxidants? O-2, 1O2, .OH, H2O2, Cu, Fe. R•, RO•, ROO • Antioxidant Prooxidant Jail R•, RO•, ROO•, 1O2, O-2, -OH, H2O2, Cu, Fe

Preventive Antioxidants Superoxide dismutase Catalase Glutathione peroxidase Singlet oxygen quencher Transition metal chelators (EDTA) Preventive antioxidants minimize the formation of initiating radicals

Superoxide dismutase Catalase Superoxide dismutase 2O2·- H2O2 O2 + H2O Glutathione Oxidase GSSG + 2H2O 2GSH Glutathione Reductase NADP+ NADPH + H+ NADP+ Reductase

Gluthione H H H O O C N C C N CH2COOH CH2 CH2 CH2 SH HC NH2 COOH

Singlet Oxygen Quenching Mechanism of Carotenes 1O2 + 1-CAROTENE 3O2 + 3-CAROTENE 3 -CAROTENE 1 -CAROTENE RADIATIONLESS

Prooxidant Activities of Transition Metals Formations of alkyl free radical by direct reaction with fats and oils. Fe 3+ + RH Fe 2+ + R + H + · Hydroperoxide decomposition to form peroxy or alkoxy radical. Fe 3+ + ROOH Fe 2+ + ROO + H + · Fe 2+ + ROOH Fe 3+ + RO - · + OH Activation of molecular oxygen for singlet oxygen formation. Fe 2+ + O Fe 3+ + O - 1 O 2 2 2

Radical Scavenging Antioxidant Vitamin C Tocopherol Quercetin Anthocyanin Radical scavenging antioxidants break free radical chain reaction by donating hydrogen to free radicals

Standard One-Electron Reduction Potential Compounds E (mV) HO· H+ / H2O 2310 RO· H+ / ROH 1600 HOO. H+ / ROOH 1300 ROO· H+ / ROOH 1000 R· H+ / RH 600 Catechol· H+ / Catechol 530 - Tocopheroxyl· H+ / - Tocopherol 500 Ascorbate· H+ / Ascorbate 282

Resonance Stabilization of Antioxidant Radicals H C ( C H ) 3 3 E0 = 300-500mv E0=1000mv (K= 107 M-1sec-1) + R , RO , ROO • • • RH , ROH , ROOH O C H 3 . O O . C ( C H ) C ( C H ) 3 3 3 3 O C H O C H 3 3 O O . C ( C H ) C ( C H ) 3 3 3 . O C H O C H 3 3

Minimization of Lipid Oxidation If a compound inhibits the formation of free alkyl radicals in the initiation step, or if the chemical compound interrupts the propagation of the free radical chain, the compound can delay the start or slow the chemical reaction rate of lipid oxidation. The initiation of free radical formation can be delayed by the use of metal chelating agents, singlet oxygen inhibitors, and peroxide stabilizers. The propagation of free radical chain reaction can be minimized by the donation of hydrogen from the antioxidants and the metal chelating agents.

Characteristics of Antioxidants The major antioxidants currently used in foods are monohydroxy or polyhydroxy phenol compounds with various ring substitutions. These compounds have low activation energy to donate hydrogen. The resulting antioxidant free radical does not initiate another free radical due to the stabilization of delocalization of radical electron. The resulting antioxidant free radical is not subject to rapid oxidation due to its stability. The antioxidant free radicals can also react with lipid free radicals to form stable complex compounds

Antioxidants O H C ( ) C ( H ) O Butylated Butylated Hydroxy Anisole 3 ) C ( H 3 ) O Butylated Hydroxy Anisole Butylated Hydroxy Toluene

Antioxidants Gossypol O H C O H C ( ) Propyl Gallate TBHQ C H O 3 7 3

Mechanism of Antioxidants Hydrogen donation to free radicals by antioxidants. Formation of a complex between the lipid radical and the antioxidant radical (free radical acceptor).

Reaction of antioxidants with radicals · + AH RH + A · RO + AH ROH + A · · ROO + AH ROOH + A · · R + A RA · · RO + A ROA · · ROO + A ROOA · · Antioxidant + O Oxidized Antioxidant 2

Stable Resonance Formation of BHA ( 3 ) R · , RO · , or ROO · . C ( H 3 ) O O C H 3 ( ) . RH, ROH + or ROOH C ( H 3 ) O O . C ( C H 3 ) 3 . O C H 3

Tocopherol and Oxygen Reaction 3 2 ( ) a - tocopherol C H C H C H 3 3 3 ) C H ( C H ) C H ( C H ) 3 2 3 3 2 O H a - tocoquinone

Mechanisms of Metals in Accelerating Lipid Oxidation Formations of alkyl free radical by direct reaction with fats and oils. Fe 3+ + RH Fe 2+ + R + H + · Hydroperoxide decomposition to form peroxy or alkoxy radical. Fe 3+ + ROOH Fe 2+ + ROO + H + · Fe 2+ + ROOH Fe 3+ + RO - · + OH Activation of molecular oxygen for singlet oxygen formation. Fe 2+ + O Fe 3+ + O - 1 O 2 2 2

Kinds of Metal Chelators Metal chelators deactivate trace metals that are free or salts of fatty acids by the formation of complex ion or coordination compounds. 1. Phosphoric acid 2. Citric acid 3. Ascorbic acid 4. Ethylene-Diamine-Tetra-Acetate (EDTA)

Metal Ions – EDTA Complex Formation H O 2 C C H 2 O N C H 2 M C H O N 2 C C H C H 2 2 O O C O

Synergism in Lipid Oxidation Synergism occurs when mixtures of antioxidants produce a more pronounced activity than the sum of the activities of the individual antioxidants when used separately. To have maximum efficiency, primary antioxidants are often used in combination with (1) other phenolic antioxidants, or with (2) various metal chelating agents.

Factors Affecting the Efficiency of Antioxidant 1. Activation energy of antioxidants to donate hydrogen should be low 2. Oxidation potential should be high 3. Reduction potential should be low 4. Stability to pH and processing. 5. Solubility in oil should be .

Antioxidant Safety Food Additive, Meat Inspection, and Poultry Inspection Acts. Total concentration of authorized antioxidants added singly or in combination, must not exceed 200 parts per million by weight on the basis of fat content of the food.

Possible Future Antioxidants 1. Polymeric antioxidant. 2. Antioxidant attached to the packaging materials. 3. Development of new, non-absorbable polymeric antioxidants for use in foods.

Long-Term Safety of Monomeric Antioxidants Pathological effect. Carcinogenic potential Interactions with enzymes Effects of reproduction The exact nature of the metabolism rate in man.

Isolation and Identification of Oxidation Product of 2,6-Di-(Tert-Butyl)-4-Methylphenol 3,3' ,5,5'-Tetra-Bis-( Tert-Butyl)-4,4'-Dihydoxyl-1,2-Diphenylethane O C H C H O 3,3' ,5,5'-Tetra-Bis-( Tert-Butyl)- Stillbenequinone

Ideal Antioxidants No harmful physiological effects Not contribute an objectionable flavor, odor, or color to the fat Effective in low concentration Fat-soluble Carry-through effect  No destruction during processing Readily-available Economical Not absorbable by the body

Biochemical Control of Lipid Oxidation

Biochemical Control of Lipid Oxidation in Mayonnaise Composition of Mayonnaise Composition (%) Soybean oil 77.0 Whole egg 7.0 Water 7.0 Vinegar 3.0 Egg yolk 2.0 Glucose 1.0 Fructose 1.0 Salt 0.9 Natural Flavor 0.1 100%

Glucose oxidase/catalase Reaction Mechanism. 2 Gluconic acid + 2H O 2 2 2 2 Catalase 2H O 2H O + O 2 2 2 2 The net chemical reaction is: Glucose Oxidase 2 Glucose + O2 2 Gluconic acid Catalase

Kinds of Antioxidants Natural antioxidants: 1.Tocopherols (delta>gamma>beta>alpha) 2.Nordihydroguaretic Acid (NDGA) 3.Sesamol 4.Gossypol Synthetic antioxidants: 1.Butylated Hydroxy Anisole (BHA) 2.Butylated Hydroxy Toluene (BHT) 3.Propyl Gallate (PG) 4.Tertiary Butyl Hydroquinone (TBHQ)

Choices of Antioxidants Different antioxidants show substantially different antioxidant effectiveness in different fats and oils and food systems due to different molecular structures. We should consider the following: Safety Antioxidant effectiveness Off-odor Off-color Convenience of antioxidant incorporation to foods Carry-through effect Stability to pH and food processing Availability Cost Non-adsorbable, if possible

Antioxidants for Different Food Systems A small surface-to-volume ratio – PG and TBHQ A large surface-to-volume ratio – BHA and BHT

Application of Antioxidants to Foods Direct addition of antioxidants to oil or melted fat. Addition of antioxidants to the food after they are diluted in oil. Spraying antioxidant solution in oil on the food or dipping food into antioxidant solution.