1. Anti oxidants for Cosmetics Dr. Panvipa Krisdaphong Mae Fah Luang University by Dean School of Cosmetic Science

2. Outer beauty: * Outer beauty: Anti oxidants Shining healthy hair and a clear radiant complexion. The beauty that shines from inside. Younger than your chronological age. Inner beauty: * Inner beauty: * Lasting beauty: Oral Cosmetics Topical Cosmetics Topical Cosmetics & Oral Cosmetics

3. Aging 1. Genetic level – DNA 2. Cellular level - Free radical

4. Radical Toxic-chemical Sun X-ray Pollutant Smoke Diet Drugs External Source Anti-oxidation

5. Radical Theory 1) Free radical source Immune destroy virus/bacterial Activities enzyme Energy Anti-oxidation

6. 2) Dangerous of Free radical Damage to cells Damage to tissue Change protein structure Protein synthesis error Anti-oxidation

7. Free radical Atom & molecule with one or more unpaired electron - Reactive Oxygen Species (ROS) 1.Superoxide radical (O 2 ) 2.Hydrogen peroxide ( H 2 O 2 ) 3. Hydroxy radical ( OH) **

8. 1. Superoxide radical (O 2 ) Attack in body : - Enzyme - Cell membrane : “lipid peroxidation” O 2 + Unsaturated fatty acid lipid free radical brack down to cell 2. Hydrogen peroxide (H 2 O 2 ) Attack in body : - Nucleus (attack DNA : protein cross link)

9. 3. Hydroxy radical ( OH) ** ** Most potent oxidant ** Major causes of the change we see in aging Attack in body : - Enzyme - Proteins (attack DNA : most injurious to body overall) - Carbohydrates - Lipid

10. Hydroxy radical Peroxide all Reactive Oxygen Species Hydroxy radical Peroxide Superoxide Lipid peroxidation Peroxide Nucleus DNA attack Cell membrane Mitochondria** Peroxisome Cellular proteins All reactive oxygen species (ROS) Cell

11. Hydroxy radical Peroxide all Reactive Oxygen Species Hydroxy radical Peroxide Superoxide Lipid peroxidation Peroxide Nucleus DNA attack Cell membrane Mitochondria** Peroxisome Cellular proteins All reactive oxygen species (ROS) Cell

12. Mitochondria Function : - Produce energy by oxidation. - Produce free radicals - 1º sites for free radical damage Ref : The free radical theory of aging, Nathan C. Nelson, Department of Physics, Ohio State University, Columbus, OH 43201

13. Source of oxidation

15. Topical Cosmetic Oral Cosmetic Anti oxidants for Cosmetic

16. Topical Cosmetic Pomegranate Pomegranate Giant curcuma Giant curcuma Rhubarb Rhubarb

17. Botanical name: Punica granatum L. Family name : PUNICACEAE Thai name : Tab-tim ( ทับทิม ) Chinese name: Shi Liu Pi Part use : Fruits Origin : Southwest Asia. Compound :Polyphenolic compound Pomegranate

18. Traditional used : 1. Diarrheal remedy 2. Anthelmintic remedy 1. Anti-oxidant 2. Anti-bacterial Action :

19. Antioxidant activity of pomegranate fruit extract and its anthocyanidins: delphinidin, cyanidin, and pelargonidin Method : In vitro (rat brain) Examined : ESR technique* with spin trapping (Free radical scavenging activities) (Free radical scavenging activities) Result : - Pomegranate extract can exhibited scavenging activity against. OH and O 2. -. - Anthocyanidins scavenged O 2 ·- in a dose-dependent - Anthocyanidins scavenged O 2 ·- in a dose-dependent - The ID 50 values** of delphinidin, cyanidin, and pelargonidin were 2.4, 22, and 456 ų M - The ID 50 values** of delphinidin, cyanidin, and pelargonidin were 2.4, 22, and 456 ų M *ESR = Electron Spin Resonance ( found that superoxide anion free radicals in mitochondria ) ** ID 50 values = dose of inhibition Ref : Noda Y, Kaneyuki T, Morl A and Packer L., Antioxidant activity of pomegranate fruit extract and its anthocyanidins: delphinidin, cyanidin, and pelargonidin. J Agric Food Chem. 2002 Jan 2 ; 50(1) : 166-71. Antioxidant activity

20. Pomegranate (Punica granatum L.) fruits are widely consumed as juice (PJ). The potent antioxidant and anti-atherosclerotic activities of PJ are attributed to its polyphenols including punicalagin, the major fruit ellagitannin, and ellagic acid (EA) Sample Antioxidant activity Sample Antioxidant activity Pomegranate Juice ++++ Total Pomegranate Tannin +++ Punicalagin ++ Ellagic acid + Antioxidant activity In vitro antiproliferative, apoptotic and antioxidant activities of punicalagin, ellagic acid and a total pomegranate tannin extract are enhanced in combination with other polyphenols as found in pomegranate juice. Seeram NP, Adams LS, Henning SM, Niu Y, Zhang Y, Nair MG, Heber D. Center for Human Nutrition, David Geffen School of Medicine, University of California, Los Angeles, CA 90095, USA. nseeram@mednet.ucla.edu

21. Successive petroleum ether, chloroform, methanol and waterextracts of Punica granatum Ž. were tested in vitro for their antibacterial activity. Method: In vitro (Agar dilution) Test on: Test on: Staphylococcus aureus MTCC 737 Escherichia coli MTCC 723 Klebsiella pneumoniae MTCC 109 Proteus ulgaris MTCC 1771 Bacillus subtilis MTCC 441 Salmonella typhi MTCC 537 Ref : Antibacterial activity of Punica granatum D. Prashanth, M.K. Asha, A. Amit Microbiology Laboratory, Research & Deelopment Centre, Natural Remedies Pt. Ltd., Plot No. 5B, Veerasandra Indl. Area, Hosur Road, Bangalore 561 229, India Antibacterial activity

22. Result : Antibacterial activity of extractives from Punica granatum fruit * Tested material Minimum inhibitory concentration (mg/ml) Conclusion : All the tested extracts showed some antibacterial activities which were significant for the methanolic extract, especially against P. ulgaris** and B.subtilis***. ** P. ulgaris : (urinary tract infections, infant diarrhea, respiratory infections) *** B.subtilis : A common soil bacterium ( illness causing diarrhea, nausea, vomiting ) S. aureus E. coli K. pneumoniae P. ulgaris B. subtilis S. typhi Petroleum ether extract 12 12 12 12 12 12 Chloroform extract 12 12 25 12 612 Methanol extract 6 12 12 1.5 612 Water extract 25 50 Not active 12 25 25 Chloramphenicol (Standard) 0.0025 0.005 0.005 0.005 0.005 0.0025 *All determinations were done in triplicate.

23. Ref: Pomegranate juice flavonoids inhibit low-density lipoprotein oxidation and cardiovascular diseases: studies in atherosclerotic mice and in humans. Aviram M, Dornfeld L, Kaplan M, Coleman R, Gaitini D, Nitecki S, Hofman A,Aviram MDornfeld LKaplan MColeman RGaitini DNitecki SHofman A Rosenblat MRosenblat M, Volkova N, Presser D, Attias J, Hayek T, Fuhrman B.Lipid Research Laboratory, Technion Faculty ofVolkova NPresser DAttias JHayek TFuhrman B Medicine, Rappaport Family Institute for Research in the Medical Sciences, Rambam Medical Center, Haifa, Israel, 31096. aviram@tx.technion.ac.il Cardiovascular Method: in vitro - Effect of Pomegranate on macrophage oxidative status. - Effect of Pomegranate on macrophage oxidative status. - Macrophages were incubated without (control) or with - Macrophages were incubated without (control) or with increasing conc. of pomegranate for 90 min at 37 C. increasing conc. of pomegranate for 90 min at 37 C. Measurement : Cellular oxidative stress following cell incubation. - Cellular fluorescence was determined (cytometry analysis) - Cellular fluorescence was determined (cytometry analysis) As cellular oxidative stress influences both scavenger receptor- mediated uptake of Ox-LDL and cellular cholesterol biosynthesis. Result : Pomegranate shown to reduces cholesterol accumulation. - inhibit macrophage foam cell formation - inhibit macrophage foam cell formation - inhibit development of atherosclerotic lesions - inhibit development of atherosclerotic lesions

24. 0 12 25 50 75 Pomegranate Juice Concentration (mmol of total polyphenols/L) 300 250 200 150 100 50 0 Cellular Oxidative Stress Cellular Oxidative Stress (Mean Fluorescence Intensity) (Control) Conclusion: Demonstrates that upon in vitro incubation of macrophages with Ppomegranate, cellular uptake of Ox-LDL and cellular cholesterol biosynthesis were significantly reduced parallel to a significant reduction in cellular oxidative stress. These effects were possibly responsible for the observed reduction in foam cell formation and atherosclerotic lesion attenuation by Pomegranate consumption. Pomegranate reduces oxidative stress in macrophages

25. Application  Skin care product : - Anti aging products - Anti bacterial products

26. Rhubarb Botanical Name : Rheum Officinale Baill.. Common Name : Rhubarb Part Used : Root, Rhizomes

27. Rhubarb Originated This plant originated from the Asian such as * Thailand * China

28. ACTIVE INGREDIENTS Anthraquinones 1,8-dihydroxyanthraquinone; R 1 =H, R 2 =H Aaloe-emodin; R 1 =H, R 2 =CH 2 OH Chrysophanol; R 1 =H, R 2 =CH 3 Rhein; R 1 =H, R 2 =COOH Emodin; R 1 =H, R 2 =CH 3 Physicion; R 1 =OCH 3, R 2 =CH 3

29. The active ingredients in rhubarb are anthraquinnone such as rhein Analysis of HPLC

30. In Cosmetics The Rhubarb extract is use as whitening products

31. Antioxidant Phenolic Constituents in Root of Rheum officinale Method : in vitro Radical scarvenging activities (RSA) Radical scarvenging activities (RSA) Determine by: Sample + ABTS + (radical compound)* Detection by : Absorbance * 2,2'-azinobis (3-ethylbenzothiazoline-6-sulfonic acid) Antioxidant Ref:Cai Y, Sun M, Xing J, Corke H. Antioxidant phenolic constituents in roots of Rheum officinale and Rubia cordifolia: structure-radical scavenging activity relationships.J Agric Food Chem 2004 Dec 29;52 (26):7884-90.

32. Hydroxyanthraquinone from roots of R. officinale Hydroxyanthraquinone from roots of R. officinale Name Content (%) Radical scarvenging activities (mM) (mM) Anthraquinone glycoside 6.80.172 ± 0.003 Aloe emodin 1.50.173 ± 0.001 Rhein 1.90.174 ± 0.001 Emodin 2.60.172 ± 0.002 R. officinale Crude extract Alcoholic extract88.6 ± 6.4 Antioxidant Ref:Cai Y, Sun M, Xing J, Corke H. Antioxidant phenolic constituents in roots of Rheum officinale and Rubia cordifolia: structure-radical scavenging activity relationships.J Agric Food Chem 2004 Dec 29;52 (26):7884-90. Result

33. Method:in vitro Measurement: Determination of tyrosinase inhibitory activity was performed by the dopachrome method* using L-DOPA as the substrate. Result: Rhubarb extract had 28.03 % tyrosinase inhibition** at 0.5 mg/ml. at 0.5 mg/ml. *Dopachrome = intermidiate substances in the melanin biosynthesis **The potent tial tyrosinase inhibitor would show decrease dopachrome absorbtion Tyrosinase inhibitory activity

34. Evaluation of Rheum extract cream for whitening products 1. Ethical committee to ask for Ethical Clarance 2. Seletion subjects - Male or Female - Age 20-35 years - No history of skin disease such as eczema or psoriasis - No use the whitening products belong 6 month - No use the whitening products between the test

35. 3. Study design Dose:3 % rhubarb extract cream Often:Twice daily Duration:8 weeks * Compare with cream base * Compare with cream base 4. Determination of skin condition Before application:Melanin value After application:Melanin value (Every 2 week during the period of 8 weeks)

36. 5. The in vivo results Result:Increase in parameter L*after application Positive depigmenting effect Positive depigmenting effect Conclusion:Rhubarb creams (3 % ww) significantly decreased skin color intensity compare decreased skin color intensity compare with control with controlL*parameter - For clearity;from dark to light - If the L*parameter is high, the skin color is light.

37. “Evaluation of the color of the treated area” Chromameter® measurement

38. L : Clarity (dark to light) a : Green to Red spectrum b : Blue to Yellow spectrum These parameters are exploited through the calculation of the Individual. These parameters are exploited through the calculation of the Individual. Typological Angle (ITA), which defines the degree of skin pigmentation Typological Angle (ITA), which defines the degree of skin pigmentation The higher L parameter and the ITA are, the lighter the skin Principle “The chromameter converts colors into a digital code”

39. Calculation formulas Variations (  ) of the colorimetric parameters L*, b* and the ITA° were calculated in arbitrary units according to the following formula: Variations (  ) = Ti - T0 Ti : value of the colorimetric parameters on the treated zone on D28 and D56 Ti : value of the colorimetric parameters on the treated zone on D28 and D56 T0: value of the colorimetric parameters on the treated zone on D0 T0: value of the colorimetric parameters on the treated zone on D0

40. Oral Cosmetic Resveratrol Resveratrol Black paper Black paper Emblic Emblic beta glucan beta glucan

41. Resveratrol Source : Bread fruits Function: Dose: Anti-oxidant Anti-oxidant Anti-cancer Anti-cancer Cardioprotective Cardioprotective Potential cholesterol lowering effect Potential cholesterol lowering effect 30 - 50 mg day Compound: Polyphenolic compound

42. Cardioprotective effect Inhibition of platelet aggregation Promotion of vasodilation by enhancing the production of nitric oxide Inhibition of inflammatory enzymes Ref 1: Ray PS, Maulik G, Cordis GA, et al. The red wine antioxidant resveratrol protects isolated rat hearts from ischemia reperfusion injury. Free Rad Biol Med. 1999; 27:160-169 Ref2:Pace-Asciak CR, Hahn S, Diamandis EP, et al. The red wine phenolics trans-resveratrol and quercetin block human platelet aggregation and eicosanoid synthesis: implications for protection against coronary heart disease. Clin Chim Acta. 1995; 235:207-219.

43. Anti oxidation Antioxidant (on blood platelet function) It is a potent antioxidant, reactive oxygen species scavenger and metal chelators. Resveratrol reduces lipid peroxidation, oxidation and nitration of platelet and plasma proteins. Lipid peroxidation Nitration of platelet Plasma proteins Platelet recruitment Thrumbus formation Cardiovasculardisease Oxidation of Resveratrol (-)inhibit

44. Inhibition of platelet aggregation Method:in vitro induce platelet aggregation by collagen, thrombin, and ADP* Dose: 10-1000 µmol/l resveratrol Result Inducer % Platelete aggregation Control Resveratrol Resveratrol Resveratrol Aspirin (10 µ mol/l) (100 µ mol/l) (1000 µ mol/l) (10 µ mol/l) (10 µ mol/l) (100 µ mol/l) (1000 µ mol/l) (10 µ mol/l) Thrombin 82.9  12.5 64.0  13.9 a 43.6  6.3 b 30.6  5.2 b 52.5  4.6 b ADP 64.8  5.1 55.2  6.2 a 46.2  9.3 b 15.5  2.3 b 48.8  6.3 b Collagen 74.0  4.0 60.6  4.8 a 47.2  7.1 b 26.9  4.3 b 43.9  4.7 b a p<0.05, b p<0.01, compared with control group. *ADP : Adenosin diphosphate Ref: Effects of red wine and wine polyphenol resveratrol onplatelet aggregationin vivo and in vitro ZHIRONG WANG1, YUANZHU HUANG1, JIANGANG ZOU1, KEJIANG CAO1, YINAN XU1 and JOSEPH M. WU21Department of Cardiology, The First Affiliated Hospital of Nanjing Medical University, Nanjiang 210029, China;2Department of Biochemistry and Molecular Biology and Brander Cancer Research Institute,New York Medical College, Valhalla, NY 10595, USAReceived October 22, 2001; Accepted November 12, 2001

45. Vascular Protection Resveratrol can protecting the vascular walls from oxidation, inflammation, platelet aggregation, and thrombus formation. In this review, we focus on the mechanism of resveratrol cardiovascular benefic effects. Ref : Resveratrol: preventing properties against vascular alterations and ageing.Delmas D, Jannin B, Latruffe N. University of Burgundy, Laboratory of Molecular and Cell Biology, Dijon, France.Delmas DJannin BLatruffe N

46. Anti-cancer and Immune stimulating Preliminary test are findings of some resveratrol-related anti-cancer and immune-stimulating effects. Method:in vitro studies Conclusion:1. Resveratrol can inhibit tumor initiation, promotion and progression. 2. inhibit DNA synthesis and ribonucleotide reductase in mammalian cells.

47. Botanical name : Piper nigrum Linn. Family name : PIPERACEAE Part used : Seed Origin: Jungles of southwestern Asia Black paper

48. Antioxidant efficacy Ref : Vijayakumar RS, Surya D, Nalini N.Department of Biotechnology, Annamalai Nagar, Tamilnadu, India. Method:In vivo Subject:30 male rats. Duration:10 weeks. Result:Supplementation with black pepper or piperine lowered TBARS or piperine lowered TBARS And CD levels and maintained SOD, CAT, And CD levels and maintained SOD, CAT, GPx, and GSH levels to near those GPx, and GSH levels to near those of control rats. of control rats. Conclusion:Supplementation with black pepper or the active principle of black pepper, piperine, the active principle of black pepper, piperine, can reduce high-fat diet induced oxidative can reduce high-fat diet induced oxidative stress to the cells. stress to the cells.

49. Botanical Name : Emblica officinalis Family name : EUPHORBIACEAE Common name : Emblic Thai name: Ma-kham-pom Part use : Fruits Emblica officinalis

50. It grows in tropical and subtropical parts of China, India, Indonesia and Thailand Description Traditional medicine : Treat broad spectrum of disorders TonicTonic RejuvenativeRejuvenative AstringentAstringent AntipyreticAntipyretic DiureticDiuretic Anti-tussiveAnti-tussive ExpectorantExpectorant Anti-diarrhealAnti-diarrheal

51. Ascorbic acidAscorbic acid Flavonoids Flavonoids Tannins Tannins AlkaloidsAlkaloids DiterpeneDiterpene TriterpeneTriterpene Benzenoid (Gallic acid) Benzenoid (Gallic acid) Composition

52. In vitro In vitro - Anti oxidant - Anti inflammatory - Anti bacterial - Anti fungal - Anti viral - Anti mutagenic In vivo In vivo - Ulcer protective activity - Anti diabetic activity - Reduce lipid level - Protective liver injury - Immuno-modulatory - Anti-oxidant Scientifically published data

53. Active Compound Gallic acid Hydrolyzable tannin

54. Anti-oxidation of Emblic Environment Reactive Oxidation Species (ROS) O 2, H 2 O 2, OH Damage of the cell Free radicals Oxidative Stress Emblic extract Anti-oxidation (-) SOD, Catalase etc.

55. Ref : Sabu MC, Kunttan R. Antidiabetic activity of medical plant and its relation ship with their antioxidant activity. J Ethanopharmacol 2002;81:155-60 Activity: Anti-oxidant Method:In vitro Material:Methanolic extract Result:The concentration needed for the inhibition of hydroxyl radical scavenging were 155.5 mg/ml, and that for superoxide scavenging activity were found to be 6.5 mg/ml Scientifically published data

56. Ref : Characterizing the antioxidant activity of amla(Phyllanthus emblica) extractS. M. Khopde†, K. Indira Priyadarsini†, H. Mohan†, V. B. Gawandi†,J. G. Satav‡, J. V. Yakhmi†, M. M. Banavaliker**, M. K. Biyani** and J. P. Mittal†,*,#Chemistry Group, and ‡Radiation Biology Division, Bhabha Atomic Research Centre, Trombay, Mumbai 400 085, India**Ajanta Pharma Ltd, Kandivali, Mumbai 400 067, India#Also with the Jawaharlal Nehru Centre for Advanced Scientific Research, Bangalore 560 064, India Activity: Anti-oxidant Method:In vivo Test on :rat liver Dose:5-500 mg/ml Result: Emblic extracts are able to fully suppress tumer cell after 4 days. The antioxidant activity of the extract was found to be concentration-dependent. Scientifically published data

57. Antioxidant Activity Method : In vitro (DPPH method) Determined : free radical scavenging by measuring antioxidant activity (DPPH) activity (DPPH) Detection : UV Spectrophotometer and ESR Spectrometer Sample : 3 fraction3 of E. offcinalis fruit extract, gallic acid and ascorbic acid and ascorbic acid The ability of three fractions of E. offcinalis fruit extract, gallic acid and ascorbic acid for antioxidant activity

58. In vitro Antioxidant Activity in Emblica offcinalis Extract against DPPH and measured by UV Spectrophotometer E. officinalis hexane extract 617.84  g/ml E. officinalis ethyl acetate extract 2.07  g/ml E.officinalis methanol extract 3.82  g/ml Ascorbic acid4.34  g/m Gallic acid2.48  g/ml Compound IC 50 Result (antioxidation)

59. In vitro Antioxidant Activity in Emblica offcinalis Extract against DPPH and measured by ESR Spectrometer Extract against DPPH and measured by ESR Spectrometer E. officinalis hexane extract 750.58  g/ml E. officinalis ethyl acetate extract 2.66  g/ml E.officinalis methanol extract 6.02  g/ml Ascorbic acid 8.02  g/m Gallic acid 3.67  g/ml Compound IC 50 Result (antioxidation)

60. Conclusion (antioxidation) Three fractions of E. officinalis extract, gallic acid and ascorbic acid showed antioxidant activity against free radical (DPPH) both in measure by UV Spectrophotometer and ESR spectrometer. The antioxidant activity. Ethyl acetate ext.  gallic acid  methanol ext.  ascorbic acid  hexane ext.

61. Skin lightening Evaluation of Emblica officinalis extract cream Method : In vivo Subjects : 20 subjects (20-60 yr.) Instrument : Chromameter CR 300 Sample : 3 % Emblica officinalis extract cream The clinical efficacy of E. officinalis formulations were evaluated in a placebo-controlled trial using cream base as control.

62. Result The facial skin conditions were evaluated pre and post application, determined L- value. E. officinalis extract at 3 % w/w showed lightening effect start at 4 th week. effect start at 4 th week. L-value L*parameter - For clearity;from dark to light (If the L*parameter is high, the skin color is light.)

63. -Beta glucan is polysaccharide derived from the cell wall of bacterial, fungal, yeasts and cereals. - Beta glucan has many structure depend on source. Beta -glucan

64. “Beta glucan of yeast (Saccharomyces serevisae) is mainly of 1,3/1,6-β-D-glucose” Structure of Beta-glucan

65. Application Immunostimulation Cholesterol reduction Vaccine adjuvants Cosmetics for skin repair Improving food products Feed supplements

66. Application Anti oxidantion Anti inflammation

67. Application of beta glucan for Immunostimulation

69. Effect of oral administration b-glucan on cytokines JANA (2002) Application of beta glucan for Immunostimulation 0 200 400 600 800 1000 1200 IL-2IFN-gTNF-a Cytokine Cytokine level (pg/ml) Control beta-glucan - 6-wk-old BALA/c mice. - Tumor cells were injected into abdominal wall. - Adding beta-glucan (28.4 mg/kg) for 21 days. IL-2 (Interleukin 2 ) IFN-  (Interferon-γ) IFN-  (Interferon – 

70. Mean changes in ratio of total to HDL-cholesterol concentrations during supplementation with beta-glucan Am J Cli Nutr (1999) 128Baseline7 Time (wk) Application of beta glucan for cholesterol reduction 15 obese men ages 20 to 60 with high cholesterol Supplement with 7.5 g of beta-glucan in orange juice twin daily for eight weeks

71. Application of beta glucan for Cosmetics for skin repair Davis (1992 )and Donzis (1993) were used beta glucan that purified from yeast cell to cosmetic composition. The products were reduced skin irritation, lesions, cracks and dryness. Greene (2001) was applied beta-glucan to cosmetic to for the protection of skin from damage caused by ultraviolet radiation.

72. Application of beta glucan for improving food products New value products to improve health (Wheatcroft et al., 2002) - Yoghurt - Fruit juice - Salad dressing - Many food products Improving propertied food products (Thammakiti et al., 2004) - Thickening - Water-holding agent - Emulsifying stabilizer - Fat replacer