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Optimization of extraction conditions of some polyphenolic compounds from

parsley leaves (Petroselinum crispum)

Article  in  Acta Scientiarum Polonorum, Technologia Alimentaria · April 2014

DOI: 10.17306/J.AFS.2014.2.4 · Source: PubMed

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 UM P O L O
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Acta Sci. Pol., Technol. Aliment. 13(2) 2014, 145-154
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pISSN 1644-0730 eISSN 1889-9594 www.food.actapol.net/

OPTIMIZATION OF EXTRACTION CONDITIONS OF SOME

POLYPHENOLIC COMPOUNDS FROM PARSLEY LEAVES
(PETROSELINUM CRISPUM)
Paula Kuźma, Beata Drużyńska, Mieczysław Obiedziński
Department of Biotechnology, Microbiology and Food Evaluation, Warsaw University of Life Sciences – SGGW
Nowoursynowska 159, 02-787 Warsaw, Poland

ABSTRACT
Background. Parsley leaf is a rich source of natural antioxidants, which serve a lot of functions in human
body and prevent food from oxidation processes. The aim of the study was to investigate the influence of
different extraction solvents and times of extraction on natural antioxidants content. Owing to the knowledge
of the properties of extracted components and solvents, as well as their interactions, it is possible to achieve
a high effectiveness of active compounds recovery.
Material and methods. Three different extraction solvents (acetone 70% in water, methanol 80% in water
and distilled water) and different times of extraction (30 and 60 minutes) were used to determine the effi-
ciency of extraction of polyphenols and catechins, antioxidant activity against free radicals DPPH and ABTS
and the ability to chelate ion Fe2+ in dried parsley leaves. Other natural antioxidants contents in parsley leaves
were also determined.
Results and discussion. In this study the best extraction solvent for polyphenols was acetone 70% and for
catechins was distilled water. All extracts examined displayed the antioxidative activity, but water was the
best solvent in the method of assaying the activity against ABTS•+ and Fe2+ ions chelating capability, whereas
methanol turned out to be the least effective in this respect. Opposite results were observed in the case of de-
termining the activity against DPPH•. The prolongation of the extraction time enhanced or decreased antiradi-
cal activity in some cases. Additionally, important biologically active compounds in parsley leaves, such as
vitamin C (248.31 mg/100 g dry matter), carotenoids (31.28 mg/100 g dry matter), chlorophyll (0.185 mg/g
dry matter) were also analysed.

Key words: efficiency of extraction, antioxidant properties, natural antioxidants

INTRODUCTION

Natural antioxidants are significant food constitu- Polyphenols are chemical compounds being sec-
ents and serve a number of functions in a human body. ondary plant metabolites. They constitute the most
High contents of antioxidants may be found in fruits abundant group of natural antioxidants [Rosicka-Kacz-
and vegetables as well as in their processed products. marek 2004].
Their action is chiefly based on the neutralization of Carotenoids are natural pigments, synthesized by
free radicals and on the prevention from oxidation pro- higher plants, bacteria, algae and fungi [Mosiewicz
cesses [Sikora et al. 2008]. 2002]. As antioxidants, they contribute to organism


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© Copyright by Wydawnictwo Uniwersytetu Przyrodniczego w Poznaniu

Kuźma P., Drużyńska B., Obiedziński M., 2014. Optimization of extraction conditions of some polyphenolic compounds from pars-
ley leaves (Petroselinum crispum). Acta Sci. Pol., Technol. Aliment. 13(2), 145-154.

defense against damages induced by the action of the catechins and antioxidant activity of parsley leaves
reactive oxygen species [Szpetnar and Pasternak 2003]. in this study. Different times of extraction (30 min and
Vitamin C occurs in two forms: as L-ascorbic acid 60 min) were tested to check if the prolongation of
and in the oxidized form as L-dehydroascorbic acid. the extraction time will result in an increased extrac-
Both these forms display the same vitamin activity. tion effectiveness of polyphenolic compounds and will
Vitamin C is an indispensable food constituent. It par- enhance antiradical activity.
ticipates in, e.g., blood vessels tightening, it acceler-
ates blood coagulation and yields detoxifying effects MATERIAL AND METHODS
[Kleszczewska 2007].
In higher plants occur two types of chlorophyll: Plant material. The experimental material in this
a and b. Chlorophyll and its derivatives exert a posi- study were dried parsley leaves (Petroselinum cri-
tive effect on a human body by complexing selected spum) available in retail trade in Polish market. Dur-
chemical compounds claimed to be carcinogenic ing the test material was stored in a dry and dark place
agents. They are additionally very strong antioxidants in the original packages. Before extraction samples
[Ferruzzi and Blakeslee 2007, Zeno 2007]. were crushed in mortar to increase the contents of ex-
The most common method of isolating active com- tracted compounds.
pounds from a plant material is extraction with the use Chemicals. All reagents used in the study were of
of a solvent. Its effectiveness depends on many fac- pure analytical grade, unless otherwise specified. Ac-
tors, including: chemical composition of raw material, etone, sulphuric acid (VI), crystalline vanillin, ethyl
storage conditions, size and comminution degree of alcohol 96%, iron chloride (II) 4-hydrate, Folin and
a sample, method and parameters of extraction, and Ciocalteu reagents, anhydrous sodium carbonate, po-
the presence of interfering factors [Koffi et al. 2010]. tassium dichromate (VI), hexane, trisodium phosphate,
The selection of extraction conditions is mainly based potassium dihydrogen phosphate (purity ≥ 99%), ace-
on the selection of solvent type, time and tempera- tonitrile (appropriate for HPLC), and methanol (appro-
ture of extraction [Gao and Liu 2005]. Owing to the priate for HPLC) were from POCH S.A. (Gliwice, Po-
knowledge of the properties of extracted compounds land). In turn, 2,2-diphenyl-1-picrylhydrazyl (DPPH•),
and solvents, as well as their interactions, it is pos- 2,2-azino-bis-3-ethylbenzothiazoline-6-sulphonic
sible to achieve a high effectiveness of components acid (ABTS•+), (–)epicatechin, ferrozine, gallic acid,
recovery. Polar solvents are frequently employed for anhydrous sodium sulphate, methaphosphoric acid,
the recovery of polyphenols from a plant matrix. The L-cysteine (purity ≥ 99%), standard of L (+) ascorbic
most common are organic solvents, such as methanol, acid (purity ≥ 99.7%), and potassium persulphate were
acetone, ethanol and ethyl acetate. It may be caused from Sigma-Aldrich (Poznań, Poland).
by the fact that the higher molecular weight of the ex- Preparation of extracts. In order to determine the
tractant is, the lower polarity is needed to allow the effect of two extraction periods (30 min and 60 min)
substances of similar molecular weight to be extracted and three solvents (70% aqueous solution of acetone,
easily. In addition, higher phenolic contents and an- 80% aqueous solution of methanol and distilled water)
tioxidant activity are obtained using aqueous organic on the contents of polyphenols and catechins and on the
solvents than the respective absolute organic solvents antioxidative properties of parsley leaves, 6 extractions
The researchers suggest that this is due to the polar- were performed. Extracts were prepared by weighing
ity, because by increasing the proportion of more polar on an analytical scale 15-g portions of parsley leaves
water to the solvent, the polarity of the mixture also and addition of 150 ml of the respective extractant to
increases. It is also suggested that the aqueous organic each portion. The extracts were shaken at a room tem-
solvents are able to extract both high and low polar- perature: for each extractant one extract was shaken
ity compounds [Sultana et al. 2009, Uma et al. 2010]. for 30 minutes, whereas the other one for 60 min. The
Thus, aqueous methanol (80%), aqueous acetone extracts were filtrated to conical flasks with ground-in
(70%) and distilled water were chosen as the extract- stopper and kept under cold store conditions without
ants for the determination the contents of polyphenols, the access of light for maximally up to 4 weeks.

146 www.food.actapol.net/
Kuźma P., Drużyńska B., Obiedziński M., 2014. Optimization of extraction conditions of some polyphenolic compounds from pars-
ley leaves (Petroselinum crispum). Acta Sci. Pol., Technol. Aliment. 13(2), 145-154.

Total polyphenolic content. The analysis was car- water) and 4 ml of the ABTS•+ solution. After 6 min-
ried out with the Folin-Ciocalteu’s method [Singleton utes the absorbance was measured using a spectropho-
and Rossi 1965]. 300 μl of each extract was pipetted tometer Shimadzu UV-1201V at a wavelength λ = 734
into different test tubes. To this solution 4.15 ml of nm. Antioxidant activity was calculated according to
distilled water, 500 μl of sodium carbonate solution the formula: A = [(Ac – As) / Ac] × 100%, where A –
(20%) and 50 μl of Folin-Ciocalteau were added. After antiradical activity, %, Ac – absorbance of the control
20 minutes the absorbance was measured at 700 nm us- sample, As – absorbance of the experimental sample.
ing spectrophotometer Shimadzu UV-1201V. The total The ability to chelate iron ion. The analysis was
phenolic content was expressed as gallic acid equiva- carried out using iron chloride (II) and ferrozine [Lai
lent through the calibration curve of gallic acid. et al. 2001]. Extracts and reagents were collected into
Total catechin content. The analysis was con- tubes in appropriate order: 0.165 ml of suitable extract
ducted according to the method by Swain and Hillis and 2.335 ml of suitable extractant and 0.250 ml of
[1959]. To 4 tubes marked by the letters A, B, C and 2 mM solution of iron chloride (II). After 20 minutes
D were added respectively: A – 1 ml of extract, 5 ml 0.5 ml of 5 mM ferrozine solution was added to each
of distilled water, 4 ml of vanillin reagent (1 g of crys- tube. After another 10 minutes the absorbance of the
talline vanillin in 100 ml 70% H2SO4); B – 1 ml of solutions was measured using the spectrophotometer
extract, 5 ml of distilled water, 4 ml of 70% H2SO4; Shimadzu UV-1201V at a wavelength λ = 562 nm.
C – 1 ml of suitable extractant (acetone 70%, metha- The percentage ability to chelate iron ions was de-
nol 80%, water), 5 ml of distilled water, 4 ml of vanil- termined through the calibration curve of iron chlo-
lin reagent; D (blank) – 1 ml of suitable extractant, ride (II).
5 ml of distilled water, 4 ml of 70% H2SO4. After Determination of carotenoids content. The ex-
15 minutes, the absorbance of solutions was measured traction and the analysis was carried out according to
using a spectrophotometer Shimadzu UV-1201V at the the method by Sztangret et al. [2001]. 0.250 g of dried
wavelength λ = 520 nm. The result (R) was calculated parsley leaves were weighted. Next about 1.5 g of an-
according to the formula: R = A – (B + C). The total hydrous Na2SO4 was added and the whole sample was
catechin content was expressed as (–) epicatechin con- mixed. The mixture was transferred into the flask and
tent through the calibration curve of (–) epicatechin. 25 ml of hexane was added. Then the flask was shaken
DPPH radical scavenging assay. The analysis on a shaker at room temperature for 24 h. After that,
was carried out according to the method by Song et al. 2 ml of carotenoid extract and 8 ml of hexane were
[1999]. The experimental sample was prepared by tak- added into test tube. Next the absorbance was mea-
ing 4 ml of the extract and and 1 ml of DPPH• solution. sured at 450 nm using spectrophotometer Shimadzu
The control sample was prepared by taking 4 ml of UV-1201V. The total carotenoid content was was de-
suitable extraction solvent (70% acetone, 80% metha- termined through the calibration curve (180 mg of po-
nol or water) and 1 ml of the DPPH• solution. After tassium dichromate (VI) in 500 ml of distilled water.
30 min the absorbance was measured using a spectro- 1 ml of that solution is an equivalent of 2.05 μg of
photometer Shimadzu UV-1201V at a wavelength λ = β-carotene).
517 nm. Antioxidant activity was calculated according Vitamin C content by HPLC. 2 g of parsley leaves
to the formula: A = [(Ac – As) / Ac] × 100%, where: A – were weighed on an analytical scale. Next, 80 ml of
antiradical activity, %, Ac – absorbance of the control methaphosphoric acid solution with a concentration of
sample, As – absorbance of the experimental sample. 20 g/l were added and the whole sample was mixed.
Free radical scavenging ability by the use of It was then filled up with methaphosphoric acid, the
a stable ABTS radical cation. The analysis was con- flask was wrapped in aluminum foil and shaken for
ducted following the method by Re et al. [1999]. The 30 min at a room temperature. Afterwards, the sample
experimental sample was prepared by taking 40 μl of was filtrated to a conical flask. The analysis was carried
the extract and 4 ml of the ABTS•+ solution. The con- out on an analytical column Discovery C8 (150 mm ×
trol sample was prepared by taking 40 μl of suitable 4.6 mm × 5 μm) in a reversed phase system. The mo-
extraction solvent (70% acetone, 80% methanol or bile phase was gradient. The flow rate was 0.4 ml/min,

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Kuźma P., Drużyńska B., Obiedziński M., 2014. Optimization of extraction conditions of some polyphenolic compounds from pars-
ley leaves (Petroselinum crispum). Acta Sci. Pol., Technol. Aliment. 13(2), 145-154.

whereas the injected volume was 10 μl. Detection was highest effectiveness of polyphenolic compounds ex-
conducted with an UV detector at λ = 265 nm [PN-EN traction was reported for the 70% acetone solution and
14130:2004]. The time of retention was 6.148 min. the lowest for the 80% methanolic solution (Fig. 1).
Determination of chlorophyll a and b content.
The extraction and the analysis was conducted ac-
cording to the method by Dżugan [2006]. 0.250 g of
dried parsley leaves were weighted and transferred
cylinder with a stopper. Then 10 ml of methanol were
added and the suspension was shaken thoroughly.
After that the suspension was left fot 15 min in dark
place to separate the phases. Then, 5 ml of the upper
layer was collected and filtered through filter paper
wetted with methanol. To determine the contents of
chlorophyll a and chlorophyll b, 1 ml of extract and
6 ml of methanol were added to the tube. Next the ab-
sorbance of the solution was measured using a spec-
trophotometer Shimadzu UV-1201V at two different Fig. 1. Total polyphenolic content
wavelengths λ1 = 663 nm and λ2 = 645 nm. The total
chlorophyll content of chlorophyll [C] was calculated
according to the formula: C = 8.02 × A(663) + 20.2 × The statistical analysis for two-factor experiment
A(645), mg/l, where A(663) – absorbance at a wavelength demonstrated significant effect of the extracting solu-
of λ = 663 nm; A(645) – absorbance at a wavelength of tion on polyphenols content in the extract. However,
λ = 645 nm. The content of chlorophyll a Cchl(a) and there is no important influence of the extraction time
chlorophyll b Cchl(b), mg/l, was calculated according or no interaction between two factors.
to the formulas: Cchl(a) = 12.7 × A(663) – 2.69 × A(645) According to literature data, the mean content of
and Cchl(b) = 22.9 × A(645) – 4.68 × A(663). polyphenols in raw parsley leaves accounts for 1190
mg/100 g d.m. [Al-Mamary 2002]. The highest result
Statistical analysis obtained in the reported experiment was achieved af-
All assays were performed in at least 3 replications. ter 60-minute extraction with acetone.
Mean values and standard deviations were calculated Rząca and Witrowa-Rajchert [2009] report that
using Microsoft Office Excel 2002. The statistical the content of polyphenols in dried material is af-
analysis of the two-factor experiment (ANOVA) ex- fected by the method of the drying process and that
amining the effect of two factors (factor A: extraction may be the reason of extracting smaller amounts of
period occurring at two levels of variability, factor B: polyphenols. In turn, Dragovic-Uzelac et al. [2005]
type of solvent occurring at three levels of variabil- emphasize that the total polyphenols content of plant
ity) on the effectiveness of polyphenols extractions is influenced by multiple factors including species,
at a significance level of α = 0.05 and the Pearson ripening stage, cultivation and storage conditions.
correlation coefficients were carried out with STAT- Discrepancies are also likely to occur at the stage of
GRAPHICS Plus 4.1 software. experimental analyses.
According to Uma et al. [2010], the aqueous ac-
RESULTS AND DISCUSSION etone solution is the most effective solvent for extrac-
tion of polyphenols, which has been confirmed by re-
Determination of total polyphenols sults of this study. The prolongation of extraction time
The results showed that the polyphenol content had the most impact in case of methanol solution. The
ranges from 362.93 to 723.58 mg/100 g dry mass amount of polyphenols extracted decreased. Statisti-
(d.m.) depending on the nature of the solvent and cal analysis showed no differences therefore it is more
time of extraction. Out of the solvents examined, the practical to apply shorter time of extraction.

148 www.food.actapol.net/
Kuźma P., Drużyńska B., Obiedziński M., 2014. Optimization of extraction conditions of some polyphenolic compounds from pars-
ley leaves (Petroselinum crispum). Acta Sci. Pol., Technol. Aliment. 13(2), 145-154.

Determination of total catechins methods, that might have caused a decrease in cat-
The catechin content in dry parsley leaves ranges echins content. In turn, Mika et al. [2008] observed
from 5.36 to 13.82 mg/100 g d.m. depending on the that the prolongation of the extraction time resulted
type of solvent and time of extraction. The most ef- in increased effectiveness of catechins extraction. This
fective solvent for catechins turned out to be wa- was not confirmed in our study.
ter, whereas the least effective one – 80% methanol
(Fig. 2). The content of catechins determined after Antioxidative properties
aqueous extraction was over 2 times higher than that Figure 3 depicts the percentage capability of the
assayed after extraction with methanol. extracts examined to inactivate synthetic DPPH radi-
cals. The most effective in this respect turned out to
be 80% methanolic extracts, followed by 70% ac-
etone extracts. Differences in that activity between the
30-minute and 60-minute extracts were inconsider-
able. Water appeared to be the poorest extractant, for
the percentage of radicals inactivated after 30-minute
extraction accounted for 79.44%, whereas after
60-minute extraction for less than 50%. In aqueous
extracts, in the natural environment of many biologi-
cally-active compounds, some chemical and physical
changes were likely to occur that induced the partial
degradation of polyphenolic compounds and/or other
antioxidants, which in turn resulted in the suppressed
activity of these extracts.
Fig. 2. Total catechin content It has been statistically proved that there is the in-
fluence of the type of solvent and time of extraction
on extracts’ capability for DPPH• inactivation. How-
The statistical analysis of results for two-agent ex- ever, the prolongation of extraction time resulted in
periment demonstrated a significant influence of the decrease of activity against DPPH in case of aqueous
type of solvent on the quantity of extracted catechins. extracts. For methanolic and acetone extracts differ-
As for polyphenols, analysis showed no significant ef- ences between shorter and longer extraction were rath-
fects of extraction time or the correlation between the er smaller and not important. There was no correlation
factors. between polyphenols or catechins content and activity
According to the Data Base for Flavonoids Content
of the USA Department of Agriculture, the content of
catechins in dried parsley leaves is very low or none.
In a study addressing the impact of different drying
methods on losses of flavonoids, Mohd Zainol et al.
[2009] demonstrated the loss of catechins during that
process to range from 35 to 78%.
Water, which out of the studied extractants was
a poor solvent in total polyphenols assay, turned out
to be a very good solution for catechins. This was,
however, inconsistent with findings of Perva-Uzunalic
et al. [2006], who obtained the highest effectiveness
of extraction of catechins from green tea when using
70% acetone, and the lowest one – when using water.
Yet, those authors were applying different extraction Fig. 3. Capability to inactivate DPPH radicals

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Kuźma P., Drużyńska B., Obiedziński M., 2014. Optimization of extraction conditions of some polyphenolic compounds from pars-
ley leaves (Petroselinum crispum). Acta Sci. Pol., Technol. Aliment. 13(2), 145-154.

against DPPH•. It means that these variables are inde- the factors compared. In the case of the acetone and
pendent. It seems probable that these compounds did methanol extracts, the prolongation of the extrac-
not decide on antiradical activity against DPPH•. tion time resulted in enhanced chelating capability.
The activity of methanolic and aqueous extracts The difference noted in the aqueous extract was mini-
from parsley leaves against DPPH• was investigated mal. A positive, strong correlation was noted between
by Wong and Kitts [2006]. They achieved lower re- total catechins content and activity against ABTS•+.
sults, i.e. 65% activity in the methanolic extracts and It means that these variables are dependent. The high
46% activity in the aqueous extract. Yet, it should be activity of the extracts may, to some point, be due to
emphasized that the above-cited authors were apply- the presence of catechins, for the more catechins were
ing different concentrations of solvents and differ- extracted the higher the extract’s activity was. Cybul
ent extraction methods. However, a tendency may be and Nowak [2008] report that during ABTS•+ reactions
observed that in both studies methanol was a better with flavonoids some products may be formed that will
solvent in that method than water. In turn, Woźniak be characterized by extremely strong antioxidative ef-
et al. [2009] claim that extracts’ activity against fects, reacting more rapidly with the radicals than the
DPPH• depends mainly on the type of solvent applied. native compounds occurring in the extracts. In such
The results indicate that different kinds and contents cases, results of analyses may be overestimated.
of antioxidant substances which can react with DPPH• In a research by Yildiz et al. [2008] the best ex-
are extracted from plants using various extractants. tractant for the assay of the activity against ABTS•+ in
All used solvents were polar, but their polarity was lyophilized parsley leaves was 70% methanol. It was
different. The obtained results may be related to the characterized by the highest effectiveness of extraction
polarity of extractants, which increases in the follow- out of all solvents examined, i.e. aqueous solutions of
ing order: acetone > methanol > water. methanol, ethanol and acetone in concentrations of
All extracts were characterised by a very high ca- 100, 70 and 50% (v/v), and distilled water. The applica-
pability for inactivating ABTS cation radicals (Fig. 4). tion of 70% methanol enabled achieving 100% activity
The most effective turned out to be the 60-minute against ABTS•+. Our study, however, did not confirm
aqueous extraction (99.43%) and the least one – methanol to be the best extractant, but the methanolic
the 30-minute extraction with methanol (86.22%). extracts also displayed high antiradical activity.
The values obtained point to high antiradical activity Fe(II) ions complexation capability determine the
of parsley leaves. ability of components in plant extracts to chelate the
The study demonstrated a statistically significant free iron ions. Transition metal ions such as iron are
effect of the type of solvent and extraction time on important catalysts for the generation of very reactive
the antiradical activity as well as correlations between hydroxyl radicals.
Results of the assays conducted for Fe(II) ions
complexation capability are presented in Figure 5.
Water turned out to be the best solvent in that reac-
tion, because the aqueous extracts bounded more than
95% of iron ions. The lowest chelating capability was
demonstrated for the acetone extract after 30-minute
extraction (48.36%).
Analysis demonstrated the influence of the type of
solvent on chelating capability to be statistically signifi-
cant. The effect of extraction time also turned out signif-
icant, however, the effects of water and 80% methanol
on ions complexation capability did not differ signifi-
cantly between one another, but did differ when com-
pared to acetone 70% used as a solvent. The prolonga-
Fig. 4. Capability to inactivate ABTS cation radicals tion of extraction time on ions complexation capability

150 www.food.actapol.net/
Kuźma P., Drużyńska B., Obiedziński M., 2014. Optimization of extraction conditions of some polyphenolic compounds from pars-
ley leaves (Petroselinum crispum). Acta Sci. Pol., Technol. Aliment. 13(2), 145-154.

Chen et al. [2007] emphasize that the stability of ca-

rotenoid compounds during drying depends on condi-
tions and methods of the process.
In the investigated parsley leaves the content of
ascorbic acid accounted for 237.64 mg/100 g d.m. and
that of dehydroascorbic acid – for 10.67 mg/100 g d.m.
In turn, Lisiewska and Kmiecik [1997] reported ascor-
bic acid content to reach 1485 mg/100 g d.m. of fresh
parsley leaves, whereas data provided in the Slovak
Food Composition Data Bank indicate that fresh pars-
ley leaves contain 939 mg ascorbic acid/100 g d.m.
Results achieved in our study are considerably lower
Fig. 5. Fe(II) ions complexation capability than those reported elsewhere. Worthy of notice is,
however, that ascorbic acid is susceptible to the drying
resulted in increase of chelating capacity of acetone process. Hachuła et al. [2000] report that the loss of
extracts. In case of chelating abilities of aqueous and vitamin C depends, to a great extent, on the conditions
methanolic extracts the differences between shorter of plant material drying and storage. Vitamin C losses
and longer were not important. There was no linear are additionally affected by: the presence of metal ions
correlation between content of polyphenols or cat- (i.e. copper, iron), light, water activity in the product,
echins and iron ion chelating capacity. On this basis it presence of oxygen, etc., and during drying may reach
can be concluded that the variables are independent. even up to 50% [Sokhansanj and Jayas 2007].
The ability to complex iron ions can vary widely In the reported experiment, the percentage content
depending on the type of plant. This parameter gives of dehydroascorbic acid in the total content of ascorbic
an information whether compounds found in extracts and dehydroscorbic acids reached 4.3%. It is consist-
contain potential secondary antioxidants, e.g. flavo- ent with findings by Wills et al. [1984], who reported
noids [Wong et al. 2006]. This study showed that pars- the content of dehydroascorbic acid in parsley to ac-
ley leaves are probably a good source of secondary an- count for less than 5%.
tioxidants and water is undoubtedly proper extractant. The content of chlorophyll a and b in the exam-
Yildiz et al. [2008] investigated lyophilized pars- ined material reached 0.256 mg/g d.m. According to
ley leaves capability for Fe2+ ions complexation in Lisiewska and Kmiecik [1997], the respective value
methanolic extracts and achieved complexation effec- determined in fresh parsley leaves accounted for
tiveness of 68%. It is consistent with results obtained 3.93 mg/g d.m. Chlorophyll pigments are very suscep-
in the reported experiment. In turn, Wong and Kitts tible to many factors, including e.g. temperature, pH,
[2006] reported on 54% chelating capability in metha- oxygen, light or enzymes activity. During processing
nolic extracts and 47% chelating capability in aqueous they are undergoing changes, the extent of which is
extracts. The lower results may be due to a different determined by the character of plant material and pro-
chemical composition of the experimental material as cessing conditions [Minguez-Mosquera et al. 2002].
well as to losses of antioxidants at the stage of samples It may thus be speculated that in the analyzed dried
preparation or extraction. samples, chlorophyll was subject to degradation.
The content of carotenoids in the analyzed pars- Chlorophyll a occurs in a significant excess as
ley leaves reached 31.28 mg/100 g d.m. According to compared to chlorophyll b. In the studied material, the
Ben-Amotz and Fisher [1998], the mean content of ca- a:b chlorophyll ratio reached 2.65:1, which is close to
rotenoids in parsley accounts for 45.5 mg/100 g d.m., ideal ratio. In higher plants the preponderance of chlo-
whereas according to Lisiewska and Kmiecik [1997] rophyll a, comparatively to chlorophyll b should reach
for 54.3 mg/100 g d.m. In spite of the fact that in the the values of about 3:1 or even 3:2. This is the indica-
reported study the experimental material was dried, the tor of the degree of lighting assuring an adequate plant
differences in carotenoids content were insignificant. development [Olteanu et al. 2008].

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Kuźma P., Drużyńska B., Obiedziński M., 2014. Optimization of extraction conditions of some polyphenolic compounds from pars-
ley leaves (Petroselinum crispum). Acta Sci. Pol., Technol. Aliment. 13(2), 145-154.

CONCLUSIONS of antioxidant activity of plant extracts]. Herba Pol. 1,

69-78 [in Polish].
1. Parsley leaves are a rich source of antioxidants. Dragovic-Uzelac V., Pospisil J., Levaj B., Delonga K., 2005.
The study of phenolic profiles of raw apricots and apples
2. The best extractant for polyphenols extraction
and their purees by HPLC for the evaluation of apricot
turned out to be aqueous acetone 70%, whereas for
nectars and jam authenticity. Food Chem. 91, 373-383.
catechins extraction – distilled water. Dżugan M., 2006. Czynniki wpływające na stabilność zie-
3. All extracts examined displayed the antioxida- lonych barwników roślin [Factors affecting the stability
tive activity. Distilled water was the best solvent in of the green vegetable dyes]. Zesz. Nauk. PTG 7, 27-33
the method of assaying the activity against ABTS•+ [in Polish].
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and acetone turned out to be the less effective in this and cancer preventative activity of dietary chlorophyll
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was the best extractant, whereas water was the least extraction of flavonoids from cultured cells of Saussu-
effective. rea medusa Maxim. World J. Microbiol. Biotechnol. 21,
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Hachuła U., Zawisza B., Winkler W., 2000. Zastosowanie
enhanced antiradical activity. However, a longer time
metody chromatograficzno-spektrofotometrycznej do
increases the cost of analysis. In cases where there was
analitycznej kontroli kwasu L-askorbinowego w mate-
no much difference in extracted amounts of polyphe- riale farmaceutycznym i roślinnym [Application of the
nols, catechins or in antioxidative properties, it is more chromatographic-spectrophotometric method for ana-
economical to apply shorter time of extraction. lytical control of L-ascorbic acid in pharmaceutical and
5. Apart from polyphenolic compounds, the extracts plant material]. Rocz. PZH 1, 71-78 [in Polish].
examined contained other biologically-active compounds Kleszczewska E., 2007. Biologiczne znaczenie witaminy C
with beneficial effects on a human body. High contents ze szczególnym uwzględnieniem jej znaczenia w meta-
were determined for vitamin C (248.31 mg/100 g d.m.) bolizmie skóry [The biological importance of vitamin C
and natural pigments: carotenoids (31.28 mg/100 d.m.) with particular emphasis on its role in the metabolism of
and chlorophyll (0.185 mg/g d.m.). the skin]. Pol. Merk. Lek. 23, 138, 462-465 [in Polish].
6. Further studies need to be carried out in order to Koffi E., Sea T., Dodehe Y., Soro S., 2010. Effect of solvent
type on extraction of polyphenols from twenty three Ivo-
determine what are the major contributors to the high
rian plants. J. Anim. Plant Sci. 5, 550-558.
antioxidant activity of different parsley leaf extracts.
Lai L.S., Chou S.T., Chao W.W., 2001. Studies on the anti-
oxidative activities of Hsian-tsao (Mesona procumbens
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 Kuźma P., Drużyńska B., Obiedziński M., 2014. Optimization of extraction conditions of some polyphenolic compounds from pars-
ley leaves (Petroselinum crispum). Acta Sci. Pol., Technol. Aliment. 13(2), 145-154.

OPTYMALIZACJA WARUNKÓW EKSTRAKCJI NIEKTÓRYCH ZWIĄZKÓW POLIFENOLOWYCH

Z LIŚCI PIETRUSZKI (PETROSELINUM CRISPUM)

STRESZCZENIE
Wstęp. Natka pietruszki jest bogatym źródłem naturalnych antyoksydantów, które pełnią wiele funkcji w or-
ganizmie człowieka i zapobiegają procesom utleniania w żywności. Celem pracy było zbadanie wpływu róż-
nych rozpuszczalników i czasów ekstrakcji na zawartość naturalnych antyoksydantów w liściach pietruszki.
Znajomość właściwości ekstrahowanych składników oraz ich interakcji daje możliwość osiągnięcia dużej
efektywności ekstrakcji związków bioaktywnych.
Materiał i metodyka. Trzy różne rozpuszczalniki (roztwór wodny acetonu 70%, roztwór wodny metanolu
80% i woda destylowana) oraz różne czasy ekstrakcji (30 i 60 min) były użyte do określenia efektywności
ekstrakcji polifenoli i katechin, aktywności przeciwutleniającej wobec wolnych rodników DPPH i ABTS
oraz zdolności ekstraktów do chelatowania jonów Fe2+ w suszonych liściach pietruszki.
Wyniki i wnioski. Najlepszym ekstrahentem do ekstrakcji polifenoli ogółem był roztwór wodny acetonu
o stężeniu 70%, a do ekstrakcji katechin – woda. Wszystkie ekstrakty wykazywały właściwości przeciwu-
tleniające, ale woda była najlepszym rozpuszczalnikiem w metodzie określania aktywności wobec rodnika
ABTS i chelatowania jonów żelaza, natomiast najmniej efektywny był metanol o stężeniu 80%. Określając
aktywności ekstraktów wobec rodników DPPH, zaobserwowano tendencję odwrotną. W niektórych przypad-
kach stwierdzono wzrost lub zmniejszenie aktywności antyrodnikowej wraz z wydłużeniem czasu ekstrakcji.
Dodatkowo w liściach pietruszki zidentyfikowano ważne, biologicznie aktywne związki takie, jak witami-
na C (248,31 mg/100 g suchej masy), karotenoidy (31,28 mg/100 g suchej masy), chlorofil (0,185 mg/g
suchej masy).

Słowa kluczowe: efektywność ekstrakcji, właściwości przeciwutleniające, przeciwutleniacze naturalne

Received – Przyjęto: 22.04.2013 Accepted for print – Zaakceptowano do druku: 8.01.2014

For citation – Do cytowania

Kuźma P., Drużyńska B., Obiedziński M., 2014. Optimization of extraction conditions of some polyphenolic compounds from pars-
ley leaves (Petroselinum crispum). Acta Sci. Pol., Technol. Aliment. 13(2), 145-154.

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