*Thomas Wu, B Adv Sci

Supervisor: *Dr Cheang Khoo

*School of Health and Science, University of Western Sydney, Campbelltown,
NSW, Australia
• Molecules that prevent potentially
disease producing cell damage.
• At least 8000 known naturally occurring
antioxidants.
• US$5.6 billion annually.
• Wide range of functional
properties including the
reduction in the effects of
coronary heart disease,
food preservation.
• Class of plant metabolites: one or more
aromatic rings with at least one attached –
OH.
• Used extensively as dyes, food
preservatives and health supplements.
• Reduces oxidative stress, potential anti-
inflammatory and antimutagenic
properties.
• Largest subclass of Polyphenols
• Common sources of flavonoids include
citrus fruits, berries, onions, tea and dark
chocolate (> 70%)
• Preliminary research has shown
Anti-microbial, anti-allergic,
Anti-diarrheal and
Anti-viral properties.

Quercitin
• Commonly known as Old man weed.
• Native Australian Plant found along the
Murray river and in low swampy areas.
• Traditionally used for treating wounds
and infections.
• Used in cosmetics,
Anti-inflammatory
And antioxidant.
• Concentration of 2 types of antioxidants
to be determined.
• Separate colorimetric method used for
polyphenol and flavonoid.
• Final concentrations compared to HPLC
results.
•Determined using the Folin-Ciocalteu
assay.
•Gallic acid was used as standard.
• Sample mixed with F-C reagent and
5%(W/V) Na2CO3 and incubated at 45oC
for 30min.
•Absorbances measured at 760nm.
•Determined using the Aluminium
Chloride assay.
• Quercitin used as standard.
• Sample mixed with (1:20 w/v) NaNO2,
(1:10 w/v) AlCl3, 1M NaOH and distilled
water.
•Absorbance was measured at 460nm
 Gallic Acid Calibration Cruve
0.7

0.6 y = 0.0486x
R² = 0.9704
0.5
Absorbance

0.4

0.3

0.2

0.1

0
0 2 4 6 8 10 12 14
Concentration (mg/mL)

Sample Concentration from Graph Concentration of Stock

(mg/mL) solution (mg/g)

201 (dry herb) 2.366 9.465

202 (dry herb) 2.418 9.671

203 (dry herb) 2.521 10.082

204 (liquid extract) 3.107 6.21

205 (liquid extract) 4.979 9.959

 Quercitin Calibration Curve
1
0.9
0.8
0.7 y = 0.0181x
absorbance

0.6 R² = 0.9373
0.5
0.4
0.3
0.2
0.1
0
0 10 20 30 40 50 60
concentration (mg/mL)

Sample Concentration From Graph Concentration from Stock

(mg/mL) Solution (mg/g)*

201 (dry herb) 43.619 43.619

202 (dry herb) 39.900 39.900

203 (dry herb) 50.304 50.304

204 (liquid extract) 14.061 14.061

205 (liquid extract) 15.276 15.276

 Sample Concentration of Concentration of Flavonoids/Polyphenol
polyphenol (mg/g Flavonoid (mg/g QE) (F/P ratio)
GAE)

201 (dry herb) 9.465 43.619 4.608

202 (dry herb) 9.671 39.900 4.125

203 (dry herb) 10.082 50.304 4.989

204 (liquid extract) 6.21 14.061 2.264

205 (liquid extract) 9.959 15.276 1.534

 Sample Polyphenols %difference Flavonoids %
difference

Colorimetric HPLC (-HPLC) Colorimetric HPLC (- HPLC)

201 (dry 9.465 12.7 -34.18 43.619 3.8 +1047.87

herb)

202 (dry 9.671 9.0 +7.46 39.900 3.5 +1040

herb)

203 (dry 10.082 10.0 +0.82 50.304 3.0 +1576

herb)

204 (liquid 6.21 1.70 +265.29 14.061 0.17 +8171.18

extract)

205 (liquid 9.959 1.61 +518.57 15.276 0.17 +8885.89

extract)
• Evans, C. R. (2001): Flavonoid Antioxidants. Current Medicinal Chemistry, 8, 797-807
• Shahdi, F.; Janitha, P. K.; Wanasundara, P. D. (1992): Phenolic Antioxidants. Critical Reviews in
Food Science and Nutrition, 32 (1), 67-103
• Stevenson, D. E.; Hurst, R. D. (2007): Polyphenolic phytochemicals- just antioxidants or much
more?. Cell. Mol. Life Sci, 64, 2900-2916
• Sulaiman, C. T.; Balachandran, I. (2012): Total Phenolics and Total Flavonoids in Selected Indian
Medicinal Plants. Indian J. Pharm. Sci., 74 (3), 258-260
• Hart, R. S. (2012): That 'Old Man Weed'. Research Directions UWS
• Beattie, K. D.; Waterman, P. G.; Leach, D. N. (2011): Centipeda cunninghamii, an australian
traditional medicinal plant , 12, 77- PL27
• Evans, C. R.; Miller, N. J.; Bolwell, P. G.; Bramley, P. M.; Pridham, J. B. (1995): The relative
antioxidant activities of plant-derived polyphenolic flavonoids. Free Rad. Res., 22, 375-383
• Ainsworth, E. A.; Gillespie, K. M. (2007): Estimation of total phenolic content and other
oxidation substrates in plant tissues using Folin-Ciocalteu reagent. Nature Protocols, 2, 875-877
• Singleton, V. L.; Rossi Jr, J. A. (1965): Colorimetry of Total Phenolics with Phosphomolybdic-
Phosphotungstic Acid Reagents. Am. J. Enol. Vitic, 16, 144-158
• Zhishen, J.; Mengcheng, T.; Jianming, W. (1999): The determination of flavonoid contents in
mulberry and their scavenging effects on superoxide radicals. Food Chem., 64, 555-559
• Marinova, D.; Ribarova, F.; Atanassova, M. (2005): Total Phenolics and Total Flavonoids in
Bulgarian Fruits and Vegetables. Journal of the University of Chemical Technology and Metallurgy
2005, 40, 255-260