Diganta KalitaPost Doctoral Research AssociateSan Luis Valley Research Center

Colorado State Universityhttp://www.ppb.colostate.edu

Free Radicals(OH-, O2

- ,1O2 )

Proteins Lipids DNA/RNA1. Cardiovascular diseases (CHD)2. Cancers3. Inflammatory diseases4. Respiratory diseases5. Diabetes mellitus6. Cataract8. Aging process9. Others: Parkingson’s disease,

Alzheimer’s disease etc

Why do we need“Antioxidant”

Sources of antioxidants in the diet

Purple Majesty

CO97216-1P/P

CO97222-1R/R

http://www.ppb.colostate.edu

Potatoes…. 4th major food after rice, wheat and maize

Breeding new cultivars is gaining interest…

Mountain Rose

CO99100-1RU

Masquerade

Polyphenols

Terpenoids

Important phytonutrients in fruits, vegetables and potato

flavonolsflavanols

anthocyaninsflavones

flavanonesfsoflavones

phenolic acid

Stilebenes

curcuminoids

chalcones

lignans

caffeic acidferulic acid

chlorogenic acid

flavonoidss

kaempferolquercetin

luteolinapigenin

narigeninhesperitin

cyanidinpelargoridinmalvidin

epicatechinscatechinsepigallocatechin

genisteindaidzein

LycopeneLuteinCarotene

sesquiterpenes

carotenoids

Objective 1: Comparison of Polyphenol content and antioxidant activity of potato tubers with pomegranate and blueberries

•genotoxic

CH2

CH

C

NH2

O

Acrylamide

•neuro toxic•Probably Carcinogenic

(IARC, 1994)

Acrylamidein heated

food

2002 NFA

Free amino acid

reducing sugars120 0C

Acrylamide formation in French fries

Factors affecting : Content of asparagine & reducing sugarsTime and temperature of cookingpHSurface-to-volume ratio

Amino acidsReducing sugars

phenolics

Solution….• Breeding• Adding/blanching

• Citric acid, amino acid• Metal salts (Na, Ca, V)• Phenolics

Critical role of antioxidant on acrylamide formation

Report with endogenious phenolic compounds was limitedFood Research International (2013), 54, 753-759

Antioxidant Model system Effectcaffeic acid, catechin, cinnamicacid, ferulic acid, epicatechin

aqueous no reduction

caffeic acid, ferulic acid, emulsion reductionTBHQ, BHA, BHT, ferulic acid, vitamin C

aqueous reduction

oregano phenolic extract potato 30% reductionvirgin olive oil phenolic extract potato 45 %

enhancementBamboo leaves extracts potato 74 % reduction

Objective 1: Polyphenol content and antioxidant activity of potato tubers comparison with pomegranate and blueberries

Objective 2: Role of polyphenols in Acrylamide formation in the French fries

METHODS

Measurement of Total phenolicsFCR method from methanolic extraction ( GAE/g FW)

Antioxidant activity DPPH DPPH radical-scavenging activity (%) = [( Acontrol – Asample/Acontrol)] ×100

where A is the absorbance at 515 nm.

ABTSMonitoring absorbance of ABTS•+ radical at 734 nm

ORACfluorescence conditions: excitation at 485 nm and emission at 520 nm.

ORAC value (µmol TE/g) = hkc (Ssample – Sblank)/ (STrolox – Sblank), where h is the ratio between liters of juice and grams of fruits or vegetables, k is the dilution factor, and c is the concentration of Trolox in µmol/L

sample in water

homogenization

centrifuge

aqueous layer

ice bath overnight

ethyl acetate (EA)

concentrated EA

injected to GC-MS

GC-MS Conditions

Apparatus Varian Saturn 2000RColumn CP-Sil 24CB, AgilentCarrier gas HeFlow rate 1 ml/minInjection volume 1 ulInjection temp 250 °CTemp program 50 C 1min 100 C 2min 0.5

C/min 10 min, 100 C /min to235 C

Retention time 8.091Acquisition time 21.08 minInonisation EI(70 eV)Detection SIM (Acrylamide m/z 150

& 152 internal St.153 & 155)

Internal standard

Bromination reagent(KBr, HBr, Br2 solution)

Na2S2O3

Sodium sulfate

Extraction & Analysis of Acrylamide by GC-MS method via bromination derivative

y = 5.837x + 0.058R² = 0.996

0

1

2

3

4

5

6

7

0 0.2 0.4 0.6 0.8 1 1.2

Anal

yte

Area

/IS

Area

analyte conc/Internal Standard Conc

0

5000

10000

15000

20000

0

5000

10000

15000

20000

0

5000

10000

15000

20000

0

5000

10000

15000

20000

Std m/z 150

IS m/z 153

French fry, m/Z 150

IS m/z 153

Calibration curve

Representative chromatographs

RESULTS

0

200

400

600

800

1000

1200

1400

µg G

AE /

gTotal phenolics Chlorogenic acid

Total phenolics and chlorogenic acid in potato tubers

0

2000

4000

6000

8000

10000

12000

14000to

tal

phen

olic

s, ug

GA

E/g

DW

Total phenolics in potato, blueberries and pomegranate

0

20

40

60

80

100

% D

PPH

inhi

bitio

n

050

100150200250300

AB

TS

activ

ity, µ

mol

TE

/g0

200

400

600

800

OR

AC

(hyd

roph

ilic)

µ m

ol

TE

/g

0

3

6

9

12

15

OR

AC

(lipo

phili

c) µ

mol

TE

/g

Antioxidant activities in DPPH, ABTS and ORAC assays

y = 0.005x + 29.07R² = 0.808

0

10

20

30

40

50

60

70

80

90

100

0 5000 10000 15000D

PPH

act

ivity

total phenolics

y = 0.065x - 48.26R² = 0.949

0

100

200

300

400

500

600

700

800

0 5000 10000 15000

OR

AC

µm

ol T

E/g

total phenolics

y = 0.021x - 10.26R² = 0.925

0

50

100

150

200

250

300

0 5000 10000 15000

AB

TS

µ m

olT

E/g

total phenolics

Correlation between Total phenolics and antioxidant assays

Food Serving Size TAC/serving (µ mol of TE)

ORAC ABTS

Pomegranate 30 ml 21270 7743

Blueberries (40 g) 19345 5170

Purple Majesty 173 g 13886 3946

CO97226-2R/R 173 g 5066 3559

Antioxidant capacity per serving

Red and purple fleshed potato tubers had higher levels of total phenolics than the yellow and white tubers

Red and purple fleshed potato tubers had comparable levels of antioxidant activity with blueberries and pomegranate

0

5

10

15

mg/

g FW

asparagine reducing sugars

Asparagine and reducing sugars in potato tubers

0

200

400

600

800

1000

1200

1400

1600

1800

2000

acry

lam

ide,

µg/

kgAcryalmide in French fries

Correlation of acryalmide with asparagine, reducing sugars, and phenolics

Asparagine : glucose : chlorogenic acid a) 1 : 1 : 0b) 1 : 1 : 1c) 1 : 5 : 1

Effect of chlorogenic acid

Conclusions Red and purple fleshed potato tubers had higher levels of total phenolics than the yellow and white tubers

Red and purple fleshed potato tubers had comparable levels of antioxidant activity with blueberries and pomegranate

Potato tubers with lower levels asparagine and reducing sugars, and higher levels phenolics form lower levels of acrylamide

Acknowledgement

Dr Sastry S Jayanty

Dr David G Holm

Elita Castleberry

Colter Carroll