Plant aglycones. As the quantitative determination of flavonoid

Plant phenolic compounds are the plant biochemicals
that have one or more hydroxyl groups attached to aromatic ring. 14. These
compounds are investigated due to their physiological and morphological
importance in plants. Such as, they play an active part in the defensive
mechanism response of plants against herbivores, pathogens and stress
situations. These defensive actions of secondary metabolites could represent a
further method of plant protection
9. They play a critical role in human diet and interested of their high
antioxidant activities. Previous works revealed that, plant phenolic compounds
have anti-allergenic, anti-artherogenic, anti-inflammatory and anti-microbial

Large amounts of phenolic compounds have been
found in apples. 13 The previous studies revealed that, apple skin showed
higher phenolic content than pulp extracts of apples. These divergence were due
to high levels of catechins and chlorogenic acid. 4 Phenolic compounds are
classified in many different sub-groups (simple phenolics, phenolic acids,
cinnamic acids, coumarins and flavonoids) based on the number of carbon atoms
in the molecule 14. Flavonoids are one of the most investigated class among
the phenolic compounds due to their high biological acitivities. The extraction
of the flavonoids are generaly made by chloroform, diethyl ether, ethyl acetate
or dichloromethane. Flavonoids are responsible for different colors in the
plant tissues such as blue, purple, yellow, orange and red color5. The
flavonoid family includes flavanones, flavanonols, leucoanthocyanidins,
anthocyanins, anthocyanidins, and flavones. 14.

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Apples have been
discovered as important source of polyphenols, however, the separation and classification
remained analytically difficult. The content of phenolic compounds in apples
are the flavanol monomers, quercetin, glycosides, dihydrochalcones, esters of
caffeic and p-coumaric acids with qunic acid 12.

Thin Layer
Chromotography, known as TLC is a separation and identification method to
investigate compounds. It is used in scientific area for qualitative analysis
as it is economical, simple and quick method. It provides to analyse several
samples on the same TLC plate in a short-time. There is mobile (solvent) and
stationary phase (samples as spots or lines) in TLC. Silica-gel TLC method is
the most convenient method that is used to identify the phenolic acids and
flavonoids from different plant species 11.

Flavonoids are
present in plant tissues either as sugar conjugates or as aglycones. As the
quantitative determination of flavonoid glycosides is difficult hydrolysation
is used for quantification 7.

HPLC (High Performance Liquid
Chromatography) is an analytical method which uses high pressure in order to mobilize
the sample (mobile phase) through the column filled with chromatographic
packing material, called stationary phase (SP). HPLC is a common used technique
for both separation and quantification of plant phenolic compounds. HPLC
sensitivity and detection is based on purification of plant phenolics and
pre-concentration from complex matrices of crude plant extracts 6.

In this experiment, three techniques were used
to extract and analyse the content of apple phenolic compounds. Thin Layer
Chromatography (TLC) and hydrolysis were performed to separate and identify
phenolic compounds of apple (leaves and skin) samples. High Performance Liquid
Chromatography (HPLC) was performed for the separation, identification and
quantification of the samples.