Research Paper PPT on determination of phenolic compounds in apples and processed apple products by HPLC.

1. JOURNAL OF FRUIT AND ORNAMENTAL PLANT RESEARCH
DETERMINATION OF
PHENOLIC COMPOUNDS
IN APPLES AND
PROCESSED APPLE
PRODUCTS
MSc Part I
Seat No : 14027

2. INTRODUCTION
 Fruits and vegetables are the major source of
phenolic compounds in diet.
 Dietary intake of phenolics is estimated 1gram/day.
 Many different phenolic compounds have been
identified in apples . The two main subtypes include
:
1. Flavonoids (quercetins glycosides and catechin
and epicatechin).
2. Phenolic Compounds (caffeic acid and p-coumaric
acid).
3. Dihydrochalcones are also present.

3. Quercetins glycosides are present in
the skin of apple.
Dihydrochalcones are present in the
core and seeds of apple.
Phenolic acids are present in the
cortex of apple.

4. MATERIAL AND METHOD
MATERIAL :
 Phenolics content was measured in four cultivars of
apple : ‘Jonagold’ , ‘Sampion’ , ‘Idared’ and ‘Topaz’.
 Replicate batches of these cultivars were
processed into clear juice, cloudy juice and
applesauce by industrial procedures.
 Clear apple juices were digested with Panzym MK
at 50°C or Rohapect MA Plus at 20°C and ascorbic
acid was added to the cloud juice.

5. METHOD:
HPLC method was used to separate and quantify
phenolic compounds in apples and processed apple
products.
 The apples were divided into octants and were frozen
to -25°C and then grounded up.
o 10g of grounded apples were homogenized for 1
minute with 70% methanol. The slurry was transferred
to a 50ml volumetric flask, which was then filled to the
mark with 70% methanol.
o The mixture was filtered through Whatman No. 1 filter
paper. The filtrate was stored at −18°C prior to
analysis.

6.  Samples of sauces and juices were filtered, diluted
and extracted with 70% methanol in an ultrasonic
bath for 10 minutes before injection.
o All the samples before HPLC were diluted 1:3 (v/v)
with sodium acetate buffer (Solvent A)

7. HPLC
 HPLC was carried out using an Agilent 1100 Series
HPLC System equipped with a DAD detector.
 Phenolic compounds were separated using a
Phenomenex Fusion RP column with a guard
column.
 The mobile phase used consisted of 10.2% acetic
acid in 2mM of sodium acetate (solvent A) and
Acetonitrite (solvent B).
 The flow rate was kept constant at 0.5 ml/min for a
total run time of 72 min at 25°C.
 The system was run with a gradient pattern.
 The injection volume of the sample was 20 μl.

10. RESULT
 The concentration of phenolic compounds in the
cultivars evaluated was 857 mg/kg.
 The concentration of different groups of phenolic
compounds varied widely from cultivar to cultivar.
 The cultivar with the highest level of flavonols was
‘Sampion’ (477 mg/kg).
 The cultivar with the highest level of phenolic acids
was ‘Idared’.
 The cultivar with highest level of quercetin
glycosides were ‘Jonagold’ and ‘Topaz’.

11.  During the production of applesauce, phenolics
content did not essentially change.
 During the production of cloudy juices, phenolics
content dropped by 47%.
 During the production of clear juices with Panzym
MK, phenolics content dropped by 65%.
 During the production of clear juices with Rohapect
MA Plus, phenolics content dropped by 81%.

19. CONCLUSION
 Even though the cultivars differed significantly in
terms of morphology , they all contained about the
same amount of phenolics, the most abundant of
which were flavonols.
 Apple sauces contained more phenolics than
juices.
 Natural cloudy juices contained more phenolic
compounds than clear juices.
 In the production of clear juice, phenolic
compounds were more effectively extracted when
the temperature during enzyme treatment is higher.

20. BIBLIOGRAPHY
 Journal Of Fruit And Ornamental Plant Research ;
Vol. 14 (Suppl. 2),2006.
 Dietary intake and availability of polyphenols, J
NUTR. 130; Saclbert A., Wilska-Jeszka J.,
Markowski J., 2000.
 Flavonoids and chlorogeneic acid levels in apple
fruit: characterization of variation, Awad M.A., De
Jagger A., Van Westing L.M; 2000.

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