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Chemical and antioxidant studies on Crataegus pentagyna leaves and flowers


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The aim of this study was to investigate the antioxidant potential of Crataegus pentagyna leaves and flowers in relation to the phenolic content. The total phenolic, flavonoid and proanthocyanidin contents were determined using Folin-Ciocalteu, aluminium chloride and acid butanol assays, respectively. The radical scavenging activity was evaluated by several in vitro assays: Trolox equivalent antioxidant capacity assay, superoxide anion and hydroxyl radicals scavenging assays. In addition, ferrous ion chelating activity and 15-lipoxygenase inhibition were also investigated. Both leaf and flower extracts had high total phenolic (206.94±4.86 and 184.62±1.71 mg/g, respectively), flavonoid (57.08±0.21 and 67.04±0.52 mg/g, respectively) and proanthocyanidin contents (68.92±1.81 and 97.70±3.81 mg/g, respectively), scavenged ABTS (TEAC=0.64±0.01 and 0.65±0.01 µM Trolox equivalent to 1 µg/mL extract, respectively), superoxide (EC50=0.9±0.0 mg/mL) and hydroxyl radicals (EC50=0.9±0.0 and 0.86±0.05 mg/mL, respectively), chelated ferrous ions (EC50=1.3±0.0 and 1.9±0.0 mg/mL, respectively) and inhibited 15-lipoxygenase (EC50=129.63±0.75 and 151.76±1.65 μg/mL, respectively). It is noteworthy that both extracts showed a higher superoxide anion scavenger activity than catechin (EC50=2.2±0.0 mg/mL). The results of the study indicate that Crataegus pentagyna leaves and flowers are a promising source of natural antioxidants.
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Romanian Biotechnological Letters Vol. 19, No.6, 2014
Copyright © 2014 University of Bucharest Printed in Romania. All rights reserved
Romanian Biotechnological Letters, Vol. 19, No. 6, 2014 9859
Chemical and antioxidant studies on Crataegus pentagyna leaves and flowers
Received for publication, July 18, 2014
Accepted, November 20, 2014
1 Faculty of Pharmacy, University of Medicine and Pharmacy Grigore T. Popa-Iasi,
2 Commercial Society for Medicinal Plant Research
and Processing Plantavorel-Piatra Neamt, Romania
3 Faculty of Pharmacy, Ovidius University of Constanta, Romania
4 Faculty of Medicine, University of Medicine and Pharmacy Grigore T. Popa-Iasi, Romania
5 C. I. Negoita 1st Medical Cardiology Clinic, Sf. Spiridon Emergency Clinical Hospital, Iasi,
*corresponding author: Tel.: 0746917278; Fax: +40-232-211820; e-mail:
The aim of this study was to investigate the antioxidant potential of Crataegus pentagyna leaves
and flowers in relation to the phenolic content. The total phenolic, flavonoid and proanthocyanidin
contents were determined using Folin-Ciocalteu, aluminium chloride and acid butanol assays,
respectively. The radical scavenging activity was evaluated by several in vitro assays: Trolox
equivalent antioxidant capacity assay, superoxide anion and hydroxyl radicals scavenging assays. In
addition, ferrous ion chelating activity and 15-lipoxygenase inhibition were also investigated. Both leaf
and flower extracts had high total phenolic (206.94±4.86 and 184.62±1.71 mg/g, respectively),
flavonoid (57.08±0.21 and 67.04±0.52 mg/g, respectively) and proanthocyanidin contents (68.92±1.81
and 97.70±3.81 mg/g, respectively), scavenged ABTS (TEAC=0.64±0.01 and 0.65±0.01 µM Trolox
equivalent to 1 µg/mL extract, respectively), superoxide (EC50=0.9±0.0 mg/mL) and hydroxyl radicals
(EC50=0.9±0.0 and 0.86±0.05 mg/mL, respectively), chelated ferrous ions (EC50=1.3±0.0 and 1.9±0.0
mg/mL, respectively) and inhibited 15-lipoxygenase (EC50=129.63±0.75 and 151.76±1.65 μg/mL,
respectively). It is noteworthy that both extracts showed a higher superoxide anion scavenger activity
than catechin (EC50=2.2±0.0 mg/mL). The results of the study indicate that Crataegus pentagyna leaves
and flowers are a promising source of natural antioxidants.
Keywords: Crataegus pentagyna Willd. ssp. pentagyna, polyphenols, reactive oxygen species,
scavenging activity, chelating activity, 15-lipoxygenase, cardiovascular disease
1. Introduction
The genus Crataegus (Rosaceae, hawthorn) consists of more than 280 species that are
widespread throughout the world. Crataegus monogyna Jacq. (Lindm.) and Crataegus
laevigata (Poiret) D.C. are found and used in Europe, Crataegus pinnatifida Bunge and
Crataegus cuneata Siebold & Zucc. in China, Crataegus sinaica Boiss. in Egypt and
Crataegus aronia syn. azarolus (L.) in Israel (LJUBUNCIC & al. [1], REFAAT & al. [2],
KWOK & al. [3], JURIKOVA & al. [4]).
In Europe extracts of hawthorn leaves, flowers and fruits are used as dietary supplements
and herbal medicines in the treatment of cardiovascular disorders due to their
coronarodilating, positive inotropic, hypotensive and antiarrhythmic effects. Polyphenols,
namely flavonoids and oligomeric proanthocyanidins, have been identified as active
constituents in these extracts (VERMA & al. [5]).
9860 Romanian Biotechnological Letters, Vol. 19, No. 6, 2014
Both in vitro and in vivo studies showed the complexity and unique profile of the activity
exhibited by hawthorn extracts on the cardiovascular system. The most studied extracts were
those isolated from Crataegus laevigata. Hawthorn extracts showed positive inotropic effects
due to the inhibition of the myocardial Na+/K+-ATPase. Hawthorn extracts decreased blood
pressure by inhibition of angiotensin converting enzyme, phosphodiesterase-3 and endothelin-
1 release from endothelium thus causing vasorelaxation. In addition, hawthorn extracts
activated endothelial nitric oxide synthase with a subsequent release of high levels of nitric
oxide, a vasodilating agent. It is worthy to note that hawthorn extracts showed the ability to
regulate not only high, but also low blood pressure. The mechanism of this dual activity has
not been elucidated yet. Hawthorn extracts enhanced coronary flow, increased the ability of
cardiomyocytes to consume oxygen, protected cardiomyocytes from ischemia-reperfusion
injury and prevented reperfusion-induced arrhythmia (JURIKOVA & al. [4]), VERMA & al.
[5]). The protective activity on cardiomyocytes was associated with a significant decrease in
oxidative stress (decrease of xanthine oxidase and NADPH oxidase expressions), upregulation
of anti-apoptotic proteins (Bcl-2, Hsp 70), downregulation of pro-apoptotic proteins
(cytochrome c, caspase-3), decrease in lactate dehydrogenase and creatine kinase activity
(SWAMINATHAN & al. [6]). Apart from cardiac glycosides, hawthorn extracts showed
negative chronotropic effects without causing arrhythmia. On the contrary, they exhibited
antiarrhythmic effects. Recent studies elucidated several mechanisms with possible
implications in the antiarrhythmic activity: prolongation of action potential duration, blockade
of potassium currents, decrease in the number of ventricular ectopic beats (MÜLLER & al.
[7], [8], GARJANI & al. [9]).
Crataegus pentagyna Willd. ssp. pentagyna is spread in the southern and south-eastern
regions of Romania (PARVU & al. [10]). A literature survey revealed only few biological
investigations on the leaves and flowers of Crataegus pentagyna growing in Turkey
(ÖZYÜREK & al. [11]). In this respect, the aim of this work was to evaluate the antioxidant
potential of the leaves and flowers of Crataegus pentagyna growing in Romania in relation to
the phenolic content. The present work is part of a larger study aiming to explore the
possibility of using Crataegus pentagyna leaf and flower extracts as ingredients in dietary
supplements for cardiovascular diseases and other pathological conditions associated with
oxidative stress.
2. Materials and methods
Plant material
Crataegus pentagyna leaves and flowers were collected in Tulcea district in June 2010
and were authenticated in the Department of General Plant and Animal Biology, Faculty of
Pharmacy, Ovidius University of Constanta. Before extraction, the leaves and flowers were
air-dried in dark at ambient temperature (25°C). Herbarium voucher samples (P-l-2010, P-f-
2010) are deposited in the Department of Pharmacognosy, Faculty of Pharmacy, University of
Medicine and Pharmacy Grigore T. Popa-Iasi.
Tris(hydroxymethyl)aminomethane (Tris), hydrochloric acid and ferrous chloride were
purchased from Merck (Darmstadt, Germany). 2'-Azinobis(3-ethylbenzothiazoline-6-sulfonic
acid) diammonium salt (ABTS), ethylenediaminetetraacetic acid (EDTA), pyrogallol, Folin-
Ciocalteu's phenol reagent, 3-(2-pyridyl)-5,6-diphenyl-1,2,4-triazine-4',4"-disulfonic acid
monosodium salt (ferrozine), linoleic acid, lipoxydase from soybean were from Fluka
(Steinheim, Germany). (+)-Catechin hydrate, gallic acid, sodium carbonate, iron (II) sulfate
Chemical and antioxidant studies on Crataegus pentagyna leaves and flowers
Romanian Biotechnological Letters, Vol. 19, No. 6, 2014 9861
heptahydrate, hydrogen peroxide, sodium salicylate, (R)-(+)-6-hydroxy-2,5,7,8-
tetramethylchroman-2-carboxylic acid (Trolox) were from Sigma-Aldrich (Steinheim,
Germany). Ammonium iron (III) sulfate dodecahydrate, potassium persulfate and sodium
nitrite were obtained from Riedel-de Haën (Seelze, Germany). All other chemicals were of
analytical grade.
50 g of dried and powdered plant material (leaves and flowers, respectively) were
extracted with 500 mL methanol:water (7:3, v/v) for 6 h at room temperature under
continuous stirring. The extracts were filtered under vacuum and the residues were re-
extracted twice in the same conditions. The extracts were combined, evaporated under
reduced pressure at 40°C and freeze-dried. Both extracts were kept in sealed glass tubes at -
20°C until studied.
Total phenolic content
The total phenolic content was estimated using Folin-Ciocalteu method as described
previously (WANGENSTEEN & al. [12], SINGLETON & ROSSI [13]). Briefly, each extract
(0.04 mL) was mixed with ultrapure water (3.16 mL) and Folin-Ciocalteu's phenol reagent
(0.2 mL). After 5 min, 20% sodium carbonate (0.6 mL) was added followed by vigorous
mixing. The reaction mixture was incubated for 2 h at room temperature before the
absorbance was measured at 765 nm. The total phenolic content was expressed in terms of mg
gallic acid equivalents (GAE)/g of extract.
Flavonoid content
The total flavonoid content was determined by aluminium chloride assay (OZSOY & al.
[14]). Each extract (0.25 mL) was mixed with ultrapure water (1.25 mL) and 5% sodium
nitrite (0.075 mL). After 6 min 10% aluminium chloride (0.15 mL) was added. The mixture
was incubated for another 5 min followed by the addition of 1 M sodium hydroxide (0.5 mL)
and ultrapure water (0.275 mL). After vigorous shaking the absorbance was measured
immediately at 510 nm. The total flavonoid content was expressed in terms of mg (+) catechin
equivalents/g of extract.
Proanthocyanidin content
The total proanthocyanidin content was estimated by acid butanol assay (PORTER & al.
[15]). Briefly, the reaction mixture consisted of extract (0.5 mL), n-butanol-hydrochloric acid
reagent (95:5, v/v, 3.0 mL) and ferric reagent (2% ferric ammonium sulfate dodecahydrate in
2 N hydrochloric acid, 0.1 mL). The mixture was shaken vigorously and kept in the water
bath at 95°C. After 40 min the mixture was cooled and the absorbance was recorded at 550
nm. The total proanthocyanidin content (mg cyanidin/g extract) was calculated using the
molar extinction coefficient of cyanidin ( = 17,360 L·mol-1·cm-1) (QA'DAN & al. [16]).
Trolox equivalent antioxidant capacity assay
The assay was used to evaluate the Trolox equivalent antioxidant capacity (TEAC) of
Crataegus pentagyna extracts. In brief, different concentrations of extracts or Trolox (0.02
mL) were mixed with ABTS radical cation ethanol solution (1.98 mL; A734 nm=0.70±0.02).
The decrease in absorbance at 734 nm was measured after 6 min reaction time at 30°C. (+)-
Catechin hydrate was used as positive control. For calculating TEAC values, the percentage
of absorbance decrease as a function of concentration was plotted for each extract, Trolox and
catechin. TEAC values were calculated as follows: slope of dose-response curve of extracts or
catechin/slope of dose-response curve of Trolox (RE & al. [17]).
Superoxide anion radical scavenging assay
Superoxide anion radical scavenging activity was determined by self-oxidation of 1,2,3-
phentriol (pyrogallol) assay (WANG & al. [18]). Different concentrations of each extract (0.1
9862 Romanian Biotechnological Letters, Vol. 19, No. 6, 2014
mL) were mixed with Tris-HCl buffer (0.05 M, pH 8.0) containing 1 mM EDTA (2.8 mL)
and pyrogallol (6 mM, 0.2 mL). The mixture was vortexed and the absorbance at 325 nm was
recorded every 30 s over a period of 4 min. (+)-Catechin hydrate was used as positive control.
The inhibitory activity on the self-oxidation of 1,2,3-phentriol was calculated from the
absorbance vs. time curves using the following formula: (1-slope of extract or catechin/slope
of control) × 100.
Hydroxyl radical scavenging assay
Hydroxyl radical scavenging activity was evaluated according to a described procedure
with minor changes (JEONG & al. [19]). Different concentrations of each extract (0.225 mL)
were mixed with 20 mM sodium salicylate (0.9 mL), 1.5 mM FeSO4 (0.75 mL) and 6 mM
H2O2 (0.525 mL). After 30 min incubation at 37°C, the absorbance was measured at 562 nm.
(+)-Catechin hydrate was used as positive control. Hydroxyl radical scavenging activity (%)
was calculated using the formula: (1-absorbance in the presence of extracts or
catechin/absorbance of control) × 100.
Ferrous ion chelating assay
The ability to chelate ferrous ions was measured by ferrozine assay (DINIS & al. [20],
TUNG & al. [21]). Various concentrations of each extract (0.4 mL) were mixed with
methanol (1.48 mL), 2 mM ferrous chloride (0.04 mL) and 5 mM ferrozine (0.08 mL). After
vigorous shaking, the mixture was allowed to stand 10 min at room temperature. The
absorbance was measured at 562 nm. EDTA was used as positive control. The ferrous ion
chelating activity (%) was calculated using the equation: (1-absorbance in the presence of
extracts or EDTA/absorbance of control) × 100.
15-Lipoxygenase inhibition assay
Briefly, different concentrations of extracts (0.05 mL) were mixed with 0.2 M borate
buffer at pH 9.0 containing 10,000 U/mL of 15-lipoxygenase (0.95 mL). After 10 min
incubation at 25°C, linoleic acid in borate buffer was added (2 mL). The increase in
absorbance at 234 nm was recorded for 90 s. (+)-Catechin hydrate was used as positive
control. 15-Lipoxygenase inhibition (%) was determined on the basis of the absorbances of
the control and samples after 30 and 90 s reaction time using the formula: 100 [(Acontrol,90-
Acontrol,30)-(Asample,90-Asample,30)/(Acontrol,90-Acontrol,30)] (WANGENSTEEN & al. [12], CRETU &
al. [22]).
Statistical analysis
All experiments were performed in triplicate and all data were presented as mean values
± standard deviations. The EC50 values were calculated by linear interpolation between values
above and below 50% activity.
3. Results and discussions
Contents of total phenols, flavonoids and proanthocyanidins
Previous studies have shown that polyphenols, mainly flavonoids and proanthocyanidins,
are the major bioactive compounds in hawthorn species. They are responsible for many of the
pharmacological effects of hawthorn extracts (antioxidant, cardiovascular, anti-inflammatory
effects) (REFAAT & al. [2], VERMA & al. [5]). Therefore, the total phenolic, flavonoid and
proanthocyanidin contents in Crataegus pentagyna extracts were estimated according to the
methods described by SINGLETON & ROSSI [13], OZSOY & al. [14] and PORTER & al.
[15], respectively. These three methods are simple, rapid and inexpensive. Even if these
methods have a limited specificity, they are routinely used for the quantitative estimation of
the above mentioned phytochemicals and give accurate results (SINGLETON & ROSSI [13],
OZSOY & al. [14], PORTER & al. [15], KELM & al. [23], BLAINSKI & al. [24]). The total
Chemical and antioxidant studies on Crataegus pentagyna leaves and flowers
Romanian Biotechnological Letters, Vol. 19, No. 6, 2014 9863
phenolic content was higher in leaf extract (206.94±4.86 mg/g) in comparison to flower
extract (184.62±1.71 mg/g). Flower extract had a higher flavonoid content than leaf extract
(67.04±0.52 and 57.08±0.21 mg/g, respectively). Likewise, proanthocyanidin content,
calculated as cyanidin equivalents, was higher in flower extract (97.70±3.81 mg/g) in
comparison to leaf extract (68.92±1.81 mg/g) (Table 1).
Table 1. Total phenolic, flavonoid and proanthocyanidin contents in Crataegus pentagyna
Extract Total phenolic content
(mg gallic acid/g extract)
Flavonoid content
(mg (+)-catechin/g extract)
(mg cyanidin/g extract)
Leaf extract 206.94 ± 4.86 57.08 ± 0.21 68.92 ± 1.81
Flower extract 184.62 ± 1.71 67.04 ± 0.52 97.70 ± 3.81
Several studies have reported total phenolic contents ranging from 108.65 to 343.54 mg/g
in different extracts from Crataegus monogyna leaves with flowers (ÖZTÜRK & al. [25]),
total phenolic and flavonoid contents of 38.25-396.04 and 2.12-32.62 mg/g, respectively in
Crataegus azarolus leaf extracts (AMEL & al. [26]). In view of these literature data, it may be
concluded that both Crataegus pentagyna extracts contain significant amounts of phenolic
compounds and therefore they are expected to have strong antioxidant activity.
Free radical scavenging activity
Free radical scavenging ability of Crataegus pentagyna extracts was initially evaluated
by TEAC assay. Flower and leaf extracts showed similar TEAC values (0.65±0.01 and
0.64±0.01 µM Trolox equivalent to 1 µg/mL extract, respectively). TEAC value of catechin
was higher (5.46±0.16 µM Trolox equivalent to 1 µg/mL catechin) suggesting a stronger
antioxidant capacity (Table 2). Free radical scavenging effects of Crataegus pentagyna
extracts were further evaluated against superoxide anion and hydroxyl radicals, reactive
oxygen species involved in the pathogenesis of many cardiovascular disorders.
Oxidative stress plays an important role in the development and progression of many
cardiovascular diseases (ischemia, atherosclerosis, cardiomyopathy, cardiac hypertrophy,
congestive heart failure, hypertension) and cardiovascular complications of diabetes mellitus
(pheripheral and coronary artery diseases, diabetic retinopathy) (PHAM-HUY & al. [27],
BADESCU & al. [28]).
Reperfusion of the ischemic myocardial regions is the only way to restore normal blood
flow and prevent myocardial infarction. The increase in blood flow is associated with an
increase of oxygen level in the myocardial tissue, generation of reactive oxygen species and
oxidative stress-induced damage of myocardial tissue (ischemia-reperfusion injury) (MIMIĆ-
OKA & al. [29]). Generation of superoxide anion radical is a key event in ischemia-
reperfusion injury. It is mainly produced by xanthine oxidase, impaired mitochondrial activity
and neutrophil activity. Superoxide anion radical and other reactive oxygen species (hydrogen
peroxide, hydroxyl radical) produce oxidative damage to cardiomyocytes and endothelial cells
with an increase in vascular permeability (GALVEZ & al. [30]).
Superoxide anion radical is indirectly involved in the development of atherosclerosis; it
reduces ferric ions to ferrous ions, the latter generating hydroxyl radical via Fenton reaction.
Hydroxyl radical oxidizes polyunsaturated fatty acids in low-density lipoproteins, a process
which initiates the formation of atherosclerotique plaques. In addition, the final products of
lipid peroxidation (4-hydroxy-nonenal) are cytotoxic leading to tissue injury (GALVEZ & al.
9864 Romanian Biotechnological Letters, Vol. 19, No. 6, 2014
In hypertension, circulating leukocytes produce high levels of superoxide anion radical
and hydrogen peroxide. Superoxide anion radical converts the vasodilating nitric oxide into
the highly cytotoxic peroxynitrite anion. The decrease in nitric oxide levels causes arteriolar
vasoconstriction. Lipid peroxidation is enhanced leading to final products with
vasoconstrictor effects such as F2-isoprostanes. Reactive oxygen species promote other events
associated with hypertension (proliferation of vascular smooth muscle cells, deposition of
collagen on arterial wall) (MIMIĆ-OKA & al. [29], GROSSMAN [31]).
Reactive oxygen species induce myocardial damage resulting in cardiomyopathy.
Besides, reactive oxygen species activate enzymes (mitogen-activated protein kinases) and
transcription factors (nuclear factor-kB) which stimulate not only cardiac myocytes to
undergo hypertrophy, but also interstitial and perivascular fibrosis. Both cardiomyopathy and
cardiac hypertrophy increase significantly the risk of heart failure (MAULIK & al. [32]).
All these data clearly indicate that a reduction of oxidative stress might be beneficial in
the management of many cardiovascular diseases.
In superoxide anion radical scavenging assay, both extracts showed similar effects
(EC50=0.9±0.0 mg/mL) being more active than the positive control, catechin (EC50=2.2±0.0
mg/mL). Crataegus pentagyna extracts scavenged hydroxyl radical with comparable EC50
values (0.86±0.05 mg/mL for flower extract; 0.9±0.0 mg/mL for leaf extract); according to
the EC50 values, both extracts were only three times less active than catechin
(EC50=0.27±0.02 mg/mL) (Table 2). As catechin has been reported to be an efficient
scavenger of superoxide and hydroxyl radicals (KASHIMA, [33]), it is obvious that
Crataegus pentagyna extracts exhibited significant scavenging effects against both radicals.
Ferrous ion chelating activity
In living organisms iron exists as ferrous and ferric ions. The ferric ions are relatively
biologically inactive while ferrous ions are involved in the generation of hydroxyl radical
through Fenton reaction (VALKO & al. [34]). Therefore, ferrous ions chelation is one of the
main mechanisms of reduction of oxidative stress. Flower and leaf extracts chelated ferrous
ions with EC50 values of 1.9±0.0 and 1.3±0.0 mg/mL, respectively; EDTA, the positive
control, showed a significantly higher chelating capacity (EC50=6.46±0.25 μg/mL) (Table 2).
15-Lipoxygenase inhibition
15-Lipoxygenase plays a key role in the initiation and development of early
atheroslerotique lesions. The enzyme oxidizes polyunsaturated fatty acids in low density
lipoproteins and induces the expression of vascular cellular adhesion molecule 1 (VCAM-1)
and intercellular adhesion molecule 1 (ICAM-1) thus promoting monocyte chemotaxis and
adhesion to endothelial cells (BOLIK & al. [35]). According to the EC50 values, leaf extract
showed a higher inhibitory activity towards 15-lipoxygenase than flower extract (129.63±0.75
and 151.76±1.65 μg/mL, respectively). Both extracts were less active than the positive
control, catechin (EC50=27.66±0.57 μg/mL) (Table 2).
Table 2. Antioxidant activity of Crataegus pentagyna extracts.
Positive control
anion radical
activity (EC50)
Hydroxyl radical
Ferrous ion
Leaf extract 0.64±0.01* 0.9 ± 0.0*** 0.9 ± 0.0*** 1.3 ± 0.0*** 129.63 ± 0.75**
Flower extract 0.65±0.01* 0.9 ± 0.0*** 0.86 ± 0.05*** 1.9 ± 0.0*** 151.76 ± 1.65**
(+)-Catechin 5.46±0.16* 2.2 ± 0.0*** 0.27 ± 0.02*** - 27.66 ± 0.57**
EDTA - - - 6.46 ± 0.25** -
* µM Trolox equivalent to 1 µg/mL extract/catechin; ** μg/mL; *** mg/mL
Chemical and antioxidant studies on Crataegus pentagyna leaves and flowers
Romanian Biotechnological Letters, Vol. 19, No. 6, 2014 9865
Antioxidant activity has already been reported for other Crataegus species (C.
oxyacantha, C. orientalis, C. aronia syn. azarolus, C. monogyna, C. meyeri, C. pontica). An
ethanol extract of Crataegus orientalis leaves showed good activity in diphenylpicrylhydrazyl
(DPPH) radical scavenging assay (62.91±3.40% at 10 mg/mL) and β-carotene bleaching
assay (42.37% at 1 mg/mL) (BOR & al. [36]). An aqueous extract of Crataegus aronia leaves
inhibited the oxidation of β-carotene, 2,2'-azobis(2-amidino-propan) dihydrochloride
(AAPH)-induced plasma oxidation, iron-induced lipid peroxidation in rat liver homogenate
and increased intracellular glutathione levels; in addition, the extract showed a significant
superoxide anion scavenging activity (50% at 42±6 μg/mL in xanthine-xanthine oxidase
assay) (LJUBUNCIC & al. [1]). The antioxidant activity of methanol extracts of the leaves
and flowers belonging to 14 Crataegus species growing in different regions of Turkey was
investigated by different assays: CUPRAC (cupric reducing antioxidant capacity), FRAP
(ferric reducing antioxidant power), ABTS/persulfate and Folin assays. An extract of
Crataegus pentagyna leaves proved to be the most active among other Crataegus extracts
(TEACCUPRAC=0.378±0.004, TEACFRAP=0.132±0.001, TEACABTS=0.423±0.112 and
TEACFolin=0.752±0.201 mmol TR g-1). To the best of our knowledge, this is the single report
on the antioxidant activity of Crataegus pentagyna leaves and flowers, evaluating only the
ability to reduce cupric and ferric ions, ABTS radical cation and Folin reagent (ÖZYÜREK &
al. [11]). A comparison of the results in ABTS/persulfate assay with our data is difficult due
to different experimental protocols. However, besides ABTS radical cation scavenging
activity, our study evaluated the capacity to scavenge reactive oxygen species, chelate ferrous
ions and inhibit 15-lipoxygenase. In our study, according to the EC50 values, Crataegus
pentagyna extracts proved to be remarkable superoxide anion radical scavengers but also
efficient hydroxyl radical scavengers and inhibitors of 15-lipoxygenase.
4. Conclusions
Our study demonstrated that extracts of Crataegus pentagyna leaves and flowers contain
high levels of polyphenols and possess marked antioxidant effects. These results indicate a
potential health benefit of Crataegus pentagyna leaf and flower extracts in oxidative stress-
related cardiovascular disorders. The in vivo antioxidant and cardioprotective effects of
Crataegus pentagyna extracts need to be evaluated.
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... The total phenolic (TP) content of both extracts was determined using the Folin-Ciocâlteu reagent [12]. The absorbance was measured at 765 nm using a SPECORD 210 PLUS spectrophotometer (Analytik Jena, Germany) and the TP content was calculated from the calibration curve using chlorogenic acid as standard (2 -15 µg/mL). ...
... The quantification of total flavonoid (TF) content was based on the formation of flavonoid-aluminium chloride complexes. The absorbance was recorded at 510 nm and the results were expressed in mg (+)catechin equivalents/g of dry extract [12,27]. ...
... The TPA content, expressed in mg cyanidin/g of dry extract, was calculated using the molar absorptivity of cyanidin (ε = 17.360 L / (mol × cm)) [12,21]. DPPH free radical scavenging assay The free radical scavenging activity was evaluated by detecting the decrease of the absorbance of a methanolic DPPH solution at 517 nm in the presence of the extract according to a previously described methodology [4]. ...
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The purposes of the study were the comparison between the levels of different polyphenol groups in Crataegus pentagyna (CP) leaf and Viburnum opulus (VO) fruit extracts, the assessment of their 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity and the in vitro determination of their permeability parameters. CP extract had higher total phenolic, flavonoid and proanthocyanidin contents than VO extract (901.12 ± 3.23 vs. 231.15 ± 1.53 mg chlorogenic acid equivalents/g extract, 416.68 ± 2.03 vs. 95.33 ± 0.95 mg catechin equivalents/g extract and 112.7 ± 1.21 vs. 18.40 ± 0.87 mg cyanidin/g extract, respectively), while both extracts showed promising DPPH scavenging activity (EC50 = 15.56 ± 0.03 and 47.18 ± 0.12 μg/mL, respectively). To investigate whether the sublingual or cutaneous routes are suitable for the administration of CP and VO extracts, permeation and penetration studies were carried out in Franz diffusion cells. The polyphenols of CP and VO extracts had higher transfer across the sublingual porcine membrane (71.93% and 62.65% polyphenols in the receptor compartment, respectively) than across the chicken skin membrane (54.48% and 39.42% polyphenols in the receptor compartment, respectively). The results provide evidence that these extracts can be used to develop oromucosal dosage forms.
... All other chemicals and reagents were of analytical grade. Total phenolic content was quantified using the Folin-Ciocalteu assay as previously described [96,97] with minor changes. In brief, 0.2 mL of each extract were mixed with 3 mL of distilled water and 0.2 mL of Folin-Ciocalteu reagent. ...
... Flavonoids were quantified spectrophotometrically by aluminium chloride assay as previously described [96,97] with slight modifications. A volume of 0.5 mL of each extract was mixed with distilled water (1 mL) and 5% sodium nitrite (0.075 mL) followed by subsequent addition of 0.15 mL of 10% aluminium chloride (after 6 min) and 0.5 mL of 1 M sodium hydroxide (after other 5 min). ...
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Here, we aim to improve the overall sustainability of aquaponic basil (Ocimum basilicum L.)-sturgeon (Acipenser baerii) integrated recirculating systems. We implement new AI methods for operational management together with innovative solutions for plant growth bed, consisting of Rapana venosa shells (R), considered wastes in the food processing industry. To this end, the ARIMA-supervised learning method was used to develop solutions for forecasting the growth of both fish and plant biomass, while multi-linear regression (MLR), generalized additive models (GAM), and XGBoost were used for developing black-box virtual sensors for water quality. The efficiency of the new R substrate was evaluated and compared to the consecrated light expended clay aggregate-LECA aquaponics substrate (H). Considering two different technological scenarios (A-high feed input, Blow feed input, respectively), nutrient reduction rates, plant biomass growth performance and additionally plant quality are analysed. The resulting prediction models reveal a good accuracy, with the best metrics for predicting N-NO3 concentration in technological water. Furthermore, PCA analysis reveals a high correlation between water dissolved oxygen and pH. The use of innovative R growth substrate assured better basil growth performance. Indeed, this was in terms of both average fresh weight per basil plant, with 22.59% more at AR compared to AH, 16.45% more at BR compared to BH, respectively, as well as for average leaf area (LA) with 8.36% more at AR compared to AH, 9.49% more at BR compared to BH. However, the use of R substrate revealed a lower N-NH4 and N-NO3 reduction rate in technological water, compared to H-based variants (19.58% at AR and 18.95% at BR, compared to 20.75% at AH and 26.53% at BH for N-NH4; 2.02% at AR and 4.1% at BR, compared to 3.16% at AH and 5.24% at BH for N-NO3). The concentration of Ca, K, Mg and NO3 in the basil leaf area registered the following relationship between the experimental variants: AR > AH > BR > BH. In the root area however, the NO3 were higher in H variants with low feed input. The total phenolic and flavonoid contents in basil roots and aerial parts and the antioxidant activity of the methanolic extracts of experimental variants revealed that the highest total phenolic and flavo-noid contents were found in the BH variant (0.348% and 0.169%, respectively in the roots, 0.512% and 0.019%, respectively in the aerial parts), while the methanolic extract obtained from the roots of the same variant showed the most potent antioxidant activity (89.15%). The results revealed that an analytical framework based on supervised learning can be successfully employed in various Citation: Petrea, Ș.-M.; Simionov, I.A.; Antache, A.; Nica, A.; Oprica, L.; Miron, A.; Zamfir, C.G.; Neculiță, M.; Dima, M.F.; Cristea, D.S. An Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license ( /by/4.0/). Plants 2023, 12, 540 2 of 50 technological scenarios to optimize operational management in an aquaponic basil (Ocimum basili-cum L.)-sturgeon (Acipenser baerii) integrated recirculating systems. Also, the R substrate represents a suitable alternative for replacing conventional aquaponic grow beds. This is because it offers better plant growth performance and plant quality, together with a comparable nitrogen compound reduction rate. Future studies should investigate the long-term efficiency of innovative R aquaponic growth bed. Thus, focusing on the application of the developed prediction and forecasting models developed here, on a wider range of technological scenarios.
... In the fruits, chlorogenic and caffeic acids, hyperoside, isoquercitrin and rutin were identified (Ebrahimzadeh et al., 2018;Stefanache et al., 2019;Stefanache et al., 2020). In the leaves extract, the total phenolic content was of 206.94 mg gallic acid equivalent per g of extract, the total flavonoids content of 57.08 mg (+)-catechin equivalents per g of extract and the proanthocyanidin content of 68.92 mg cyanidin per g of extract; for the flowers, the total phenolic content was of 184.62 mg gallic acid equivalent per g of extract, the total flavonoids content of 67.04 mg (+)-catechin equivalents per g of extract and the proanthocyanidin content of 97.70 mg cyanidin per g of extract (Giurescu Bedreag et al., 2014). Total phenolic content in the fruits was of 721.1 mg gallic acid equivalents per g of extract, while the total flavonoid content of 87.92 quercetin equivalents per g extract (Ebrahimzadeh et al., 2018). ...
... The extracts from leaves, flowers and fruits showed a good antioxidant activity, the leaves extract having a higher antioxidant potential in comparison with the leaves of other Crataegus species (Ebrahimzadeh et al., 2008;Giurescu Bedreag et al., 2014;Özyürek et al., 2012;Rabiei et al., 2012;Tusa et al., 2016). The fruit extract from C. pentagyna showed in vivo anti-hypoxic activity in mice (Ebrahimzadeh et al., 2018). ...
... Studies to date have shown that rosehip (Rosa canina) extracts contain phenolic acids, proanthocyanidins, tannins, flavonoids, saturated and unsaturated fatty acids, phospholipids, minerals, galactolipids, and carotenoids [4,5]. Black hawthorn (Crataegus pentagyna) leaves and flowers contain high amounts of polyphenols with a beneficial antioxidant effect for the treatment of cardiovascular disorders [6]. Crataegus monogyna is a species with a high content of polyphenols with an antioxidant role, and the plant parts can be used either individually or in a mixture as a diuretic, antispasmodic, hepatoprotective, cardiotonic, and antioxidant [7]. ...
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Plant parts of some spontaneous fruit species were analyzed by HPLC-UV method to determine their content in phenolic compounds. Buds, leaves, flowers, and fruits were harvested from 11 wild fruit species of medicinal, food, and therapeutic interest: European crab apple, European wild pear, blackthorn, dog rose, elder, dewberry, wild blackberry, cornelian cherry, red hawthorn, black hawthorn, and green strawberry. The studied species were analyzed for the presence and amount of phenolic compounds: flavonoids from the subclasses flavanols (catechin hydrate and epicatechin) and flavonols (rutin, myricetin, quercetin, and kaempferol) together with phenolic acids belonging to the hydroxybenzoic acid subclass (ellagic, gallic, syringic, vanillic, and salicylic) and hydroxycinnamic acids (caffeic, ferulic, sinapic, p-coumaric, neochlorogenic, and chlorogenic). The obtained results confirm the presence of bioactive compounds in different plant parts in wild fruit species. The variability identified from one species to another, the different compositions of phenols within the species, provides useful data on the role of these plant parts in the human body.
... The production of cannabinoid compounds in plant cells as part of the secondary metabolites is affected by environmental factors, among which light quality is one of the most important ones that affects and controls the concentration of these compounds. However, plant responses and the effect of PAR and ultraviolet light have had different results in the synthesis of pharmaceutical compounds, and if it is examined more closely, it should be said that it depends on the type of secondary metabolite and plant species (Bedreag et al., 2014). Among the light spectra whose effects on plant secondary metabolites have been investigated, blue light probably acts as a beneficial stressor, and increasing the amount of this light puts plants in a state of poor alertness, which ultimately leads to increased production of secondary metabolites as a defense mechanism. ...
Cannabis which is a medicinal and industrial plant is native to Central Asia. It has been used as a source of food, fuel, fiber, medicine, and drugs for centuries. Cannabis has valuable agronomic traits, such as being easy to cultivate and creating diversity in organic farming. From an agricultural point of view, it is a high-yielding crop and does not require much pesticide, herbicide, and fertilizer compared to other kinds of crops, and therefore will have a less negative impact on the environment. Cannabis seeds are rich in protein and oil and have long been used by humans. Cannabis sativa seed oil contains good amounts of unsaturated fatty acids, including linoleic acid and linolenic acid, which are good for human nutrition and health and reduce cholesterol and high blood pressure. Cannabis can also be considered an excellent option for the production of two important biofuels, namely, biodiesel and bioethanol. In terms of phytochemicals, this plant is very complex and it contains more than 480 different chemical compounds. Some of these compounds belong to the primary metabolites such as amino acids, fatty acids, and steroids, while compounds such as cannabinoids, flavonoids, acetylbenoids, terpenoids, lignans (phenolic amides and lignamides), and alkaloids are among the secondary metabolites produced by the valuable cannabis plant. Therefore cannabis is similar to a very strong factory in terms of phytochemicals, which has a high value in the pharmaceutical industry. Today, the positive effect of cannabis pharmaceutical compounds has been reported in studies done on the effect of cannabis on various diseases, such as cancer, multipple sclerosis (MS), and acquired immune deficiency syndrome (AIDS), which further increases the value of this plant. As a result, due to the high importance of cannabis in pharmacy, medicine, and industry, agricultural science must study the environmental factors affecting the cultivation of this valuable plant to increase its yield and efficiency of the plant. Today, light-emitting diodes are considered an influential factor in the production of agricultural products by many researchers. Over the past years, several studies have been conducted on the effect of light spectra on plant growth and development, which have well demonstrated the importance of the blue and red spectra. Consequently, finding the right combination of light spectra for better growth and development of plants, including cannabis, is under investigation to get the best performance from the plant at the most appropriate time.
... Antioxidant activity has already been reported for Crataegus species (C. pentagyna, C. aronia, C. monogyna, C. meyeri, C. pontica) [21]. However, few studies have been carried out on the genetic diversity of this genus in Iran. ...
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Background: Crataegus spp. (Hawthorn) are used for horticultural and medicinal purposes. Objectives: In the present study, the genetic relationships among the five Iranian Crataegus species were evaluated by Inter Simple Sequence Repeats (ISSR) molecular marker. Methods: PCR reactions were performed using six primers (811, 827, 834, 845, 856, and 868) and genetic similarity was calculated based on Jacquard's similarity coefficient. Results: Six primers generated 79 products in total, of which 71 were polymorphic (89.9%), with an average of 13.1 bands per primer. The percentage of polymorphic bands ranged from 77 to 100. Primer 856 produced the highest number of bands, while the lowest was generated by primer 845. The Jacquard's similarity coefficient, derived from ISSR marker analysis, ranged from 0.164 to 0.337, indicating high genetic variation among Crataegus species in Iran. Conclusion: This study provides important data for identifying species relationships and helps develop plant breeding strategies to improve the medicinal properties of this genus in the future.
... C. pentagyna (small-flowered black hawthorn) is spread around limited geographical areas in Southern and Southeastern Europe or Turkey (Prinz et al., 2007;Turkoglu et al., 2005). Our literature survey showed only few studies on C. pentagyna reporting antioxidant activities for the extracts obtained from flowers, leaves (Bedreag et al., 2014) and fruits (Rabiei et al., 2012). Antiarrhythmic properties were found for the hydroethanolic extract of C. pentagyna leaves, being attributed to quercetin and isovitexin (Pahlavan et al., 2018). ...
Ethnopharmacological relevance: Crataegus leaves, flowers and fruits have been traditionally used to improve blood circulation, numerous preclinical and clinical studies supporting the cardiovascular benefits of Crataegus preparations. In this respect, there is very limited data on Crataegus pentagyna; in addition, the chemical profile of this species is still incompletely elucidated. Aim of the study: The objective of this study was to examine the cardiovascular benefits of Crataegus pentagyna Waldst. et Kit. ex Willd. (small-flowered black hawthorn, Rosaceae) extracts (leaf, flower and fruit ethyl acetate extracts) and the underlying mechanisms. We hypothesized that C. pentagyna extracts might exert vasodilatory effects and inhibit arginase activity due, in large part, to their polyphenolic constituents. Materials and methods: C. pentagyna extracts induced-relaxation and the mechanisms involved were studied ex vivo in isolated aortic rings from Sprague-Dawley rats. The inhibitory effects on bovine liver arginase I were assessed by an in vitro assay. Metabolite profiling of C. pentagyna extracts was performed and the most endothelium- and nitric oxide synthase-dependent; flower extract additionally reduced Ca2+ entry and, to a lesser extent, Ca2+ release from the sarcoplasmic reticulum. C. pentagyna proved to be an important source of arginase inhibitors with potential benefits in endothelial dysfunction that remains to be explored.
... & Kit. ex Willd on leaf and flower extract showed 206 GAE mg/g and 184 GAEmg/g extract respectively and scavenged ABTS (TEAC 0.64 and 0.65 µmol Trolox equivalent to1 mg/ml extract respectively) [37]. ...
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The aim of this study was to investigate polyphenolic composition of different parts (leaves, flowers and fruits) of Crataegus almaatensis Pojark, an endemic plant of Kazakhstan, and compare it to a well known European Crataegus oxyacantha L. flowers. A Qual-Quant analysis was performed based on HR-MS measurements on 22 secondary metabolites: flavonoids and phenolic acids. Another goal was to evaluate the antioxidant potency of hawthorn extracts which was expressed in the total phenolic content and DPPH scavenging potency tests. Leaf extracts from C. almaatensis were found to be the most rich in metabolites and the most active in antiradical tests (IC50 value of 48 μg/ml and TPC of 218 mg/g). The weakest potential was determined for the fruit extract of this species. According to the performed principal component analysis (PCA), the fruit extracts were not correlated with other organs of the plant, and the metabolites responsible for the extracts’ differentiation were cyanidin 3-glucoside and quercetin 3-galactoside. Based on a high correlation factor, the flowers of the Kazakh species was found to be as rich in polyphenols as the European hawthorn. The results of this study indicate that C. almaatensis is a promising source of natural antioxidants.
... (Bass et al., 1983;Amer et al., 2003). Plants are a promising source of natural antioxidants used as ingredients in dietary supplements for cardiovascular disease and other pathological conditions associated with oxidative stress (Giurescu Bedreag et al., 2014). In the present study, we investigated the ROS generation. ...
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The present paper aims to evaluate the marigold inflorescence yield, chlorophyll content and chlorophyll fluorescence of Calendula officinalis L. under ecological fertilization with four different foliar fertilizers (Fylo®, Geolino Plants&Flowers®, Cropmax®, Fitokondi®). The flowers yield was increased in all fertilized plants, compared to unfertilized ones, but the content of chlorophyll pigment and chlorphyll fluorescence values were not significantly influenced.
... Phenols, powerful antioxidants, are common secondary metabolites in plants, with potential health benefits. There have been reported antibacterial, antiviral, anti-inflammatory, anti-carcinogenic and vasodilatory actions for these chemical constituents (FÎNTÎNERU & al. [3], NAGAVANI & al. [4], GIURESCU-BERDEAG & al. [5], ROBU & al. [6]). ...
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Rudbeckia hirta L. is an ornamental plant of Asteraceae family, largely used in landscaping and prevention of soil erosion. The aim of the study was to extend the uses of this species, in relation to phenolic content. Two cultivars, 'Moreno' and 'Goldilocks', were studied. Height and plant diameter, number and diameter of inflorescences and total polyphenols content were registered related to cultivar, fertilisation and aerial organs of the plants. The chemical profile was established by TLC techniques (thin layer chromatography) and the total content of polyphenols was established by Folin- Ciocalteu method. A general spectrum of these compounds was assessed by ultra-high pressure liquid chromatography on a Thermo-Fischer UltiMate 3000 system. The results indicated that plants adapted well to local conditions and inflorescences could represent a source for these compounds in both cultivars (1,146g% and 0,8924g% for 'Goldilocks' and 'Moreno' respectively). Enriched soil conditions have significant increased the polyphenols in 'Moreno' flowers (0,8924g% and 1,801g% respectively). Chlorogenic acid and hyperoside were commonly met in Rudbeckia inflorescences.
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The present study was undertaken to investigate the hypotensive and the in vitro antioxidant activities of Crataegus azarolus L. (Rosaceae) , a plant widely used as natural remedy for hypertension in folk medicine. The antioxidant potential of methanolic extract (ME) and its three fractions of Chloroform (CHE), ethyl acetate (EAE) and water (AqE) have been investigated using several assays. Total phenolic, total flavonoid and tannins contents of the extracts were estimated, whereas EAE was also subjected to analysis by different chromatographic methods. Chemical analysis of EAE revealed to the isolation of two flavonoids; new favanol, (+)-catechin (1), and a previously identified flavonol, hyperoside (2) from C. azarolus leaves. EAE extract showed the highest polyphenolic and flavonoids contents (396.04 ± 1.20 mg GAE/g of dry extract and 32.73 ± 0.03mg QE/g of dry extract) respectively. Similarly, this extract possessed the highest antioxidant activity in all antioxidant models except for its ferrous ion chelating capacity whereas AqE and ME are the most active extracts. Intravenous administration of ME and EAE decreased mean arterial blood pressure, systolic and diastolic blood pressure in anesthetized rats dose-dependently, at the dose range of 0.4 to 12 mg/kg. In conclusion, the present study supported the significant potential to use C. azarolus by-products as a source of natural antioxidants and provides scientific justification for its traditional uses as cardioprotective and antihypertensive remedy.
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Abstract A crude hydromethanolic extract from Pinus brutia bark and its fractions (diethyl ether, ethyl acetate, n-butanol, and aqueous fractions) were studied with regard to their phenolic content and antioxidant activities. The total phenolics and proanthocyanidins in each extract were quantified by spectrophotometric methods; the polyphenolic profile was analyzed by RP-HPLC-DAD-ESI-MS. All extracts were tested with regard to their ability to scavenge free radicals (ABTS radical cation, superoxide and hydroxyl radicals), reduce ferric ions, and inhibit 15-lipoxygenase. P. brutia bark extracts had high phenolic contents (303.79±7.34-448.90±1.39 mg/g). Except diethyl ether extract, all other extracts contained proanthocyanidins ranging from 225.79±3.94 to 250.40±1.44 mg/g. Several polyphenols were identified by RP-HPLC-DAD-ESI-MS: taxifolin in diethyl ether extract, a taxifolin-O-hexoside, catechin, procyanidin dimers, and trimers in ethyl acetate extract. Except diethyl ether extract, all other extracts were effective scavengers of superoxide and hydroxyl radicals (EC50=33.5±1.1-54.93±2.85 μg/mL and 0.47±0.06-0.6±0.0 mg/mL, respectively). All extracts had noticeable 15-lipoxygenase inhibitory effects (EC50=22.47±0.75-34.43±2.25 μg/mL). We conclude that P. brutia bark is very promising for the dietary supplements industry due to its high free radical scavenging and 15-lipoxygenase inhibitory effects.
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Crataegus species (Rosaceae), known as "Howthorn" have found special medicinal use for the treatment of mild heart diseases. This work aims to measure the antioxidant capacities of various Crataegus species growing in Turkey. In this study, the flowers and leaves from 52 samples belonging to 17 taxa of 14 Crataegus species naturally growing in Turkey have been investigated for their antioxidant activity/capacity. Four different methods (CUPRAC, FRAP, ABTS/Persulfate and Folin: FCR assays) were used for determination of the antioxidant capacities of the samples. The leaves and the flowers of the plants were studied separately. Samples representing the same species collected from different locations were studied separately. The results have indicated that the samples differing by some minor morphological characteristics exhibit considerably different antioxidant capacities. Among the flower samples, the most effective species was C. x sinaica Boiss. nothosubsp. sinaica and among the leaf samples C. pentagyna Waldst and Kit. ex. Willd. were the most active. Generally, C. monogyna Jacq. samples have exhibited markedly high antioxidant activity. Moreover, the species collected from Bolu district (surrounded by several forests and lakes) have shown significantly high activity regardless of the species differences among the samples.
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Limonium brasiliense is a common plant on the southern coast of Brazil. The roots are traditionally used for treatment of premenstrual syndrome, menstrual disturbances and genito-urinary infections. Pharmaceutical preparations obtained from its roots and used for these purposes were marketed in Brazil in the 1980s and 1990s. Currently, the Brazilian Drug Agency (National Health Surveillance Agency, ANVISA) has canceled the registration of these products, and their use was discontinued because of a lack of studies to characterize the plant raw material and ensure the effectiveness and safety of its use. The aim of the present study was to develop and validate an analytical method to determine the content of total polyphenols (TP) in an extract from L. brasiliense roots, by the UV/Vis spectrophotometric method. L. brasiliense roots were extracted in acetone:water (7:3, v/v-10% w/v). The crude extract was used to develop a method for TP assay. The method was validated according to national and international guidelines. The optimum conditions for analysis time, wavelength, and standard substance were 30 min, 760 nm, and pyrogallol, respectively. Under these conditions, validation by UV/Vis spectrophotometry proved the method to be linear, specific, precise, accurate, reproducible, robust, and easy to perform. This methodology complies with the requirements for analytical application and to ensure the reliability of the results.
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Free radicals and oxidants play a dual role as both toxic and beneficial compounds, since they can be either harmful or helpful to the body. They are produced either from normal cell metabolisms in situ or from external sources (pollution, cigarette smoke, radiation, medication). When an overload of free radicals cannot gradually be destroyed, their accumulation in the body generates a phenomenon called oxidative stress. This process plays a major part in the development of chronic and degenerative illness such as cancer, autoimmune disorders, aging, cataract, rheumatoid arthritis, cardiovascular and neurodegenerative diseases. The human body has several mechanisms to counteract oxidative stress by producing antioxidants, which are either naturally produced in situ, or externally supplied through foods and/or supplements. This mini-review deals with the taxonomy, the mechanisms of formation and catabolism of the free radicals, it examines their beneficial and deleterious effects on cellular activities, it highlights the potential role of the antioxidants in preventing and repairing damages caused by oxidative stress, and it discusses the antioxidant supplementation in health maintenance.
A method for the screening of antioxidant activity is reported as a decolorization assay applicable to both lipophilic and hydrophilic antioxidants, including flavonoids, hydroxycinnamates, carotenoids, and plasma antioxidants. The pre-formed radical monocation of 2,2'-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS*+) is generated by oxidation of ABTS with potassium persulfate and is reduced in the presence of such hydrogen-donating antioxidants. The influences of both the concentration of antioxidant and duration of reaction on the inhibition of the radical cation absorption are taken into account when determining the antioxidant activity. This assay clearly improves the original TEAC assay (the ferryl myoglobin/ABTS assay) for the determination of antioxidant activity in a number of ways. First, the chemistry involves the direct generation of the ABTS radical monocation with no involvement of an intermediary radical. Second, it is a decolorization assay; thus the radical cation is pre-formed prior to addition of antioxidant test systems, rather than the generation of the radical taking place continually in the presence of the antioxidant. Hence the results obtained with the improved system may not always be directly comparable with those obtained using the original TEAC assay. Third, it is applicable to both aqueous and lipophilic systems.
The benefits of the polyphenols extracted from the seeds of black grapes, have been shown by using as experimental model the streptozotocin-induced diabetes mellitus on the Wistar rats. The streptozotocin was administered in a single dose of 60 mg/Kg body mass, intraperitoneal. The vegetal polyphenols were administered under the form of water solution, in a dose of 0,028 g/KG body mass, p.o. (through tube feeding), every two days, for a period of 16 weeks (3 weeks before the streptozotocin induction and 13 weeks after). The results obtained point out a significant improvement (p>0,001) of the lipid profile and a reduced atherogenic risk (the medium value of the atherogenity index obtained at the diabetic rats with polyphenolic protection was of 2.67 in comparison with the diabetic rats, that had an index of atherogenity of 6.23) and an insignificant improvement of the hyperglycemia (p<0.05). All the animals suffering from DM experienced various degrees of retinal detachment. Polyphenols intake by diabetic animals resulted into a low retina disease rate. They experienced only random and uneven retina detachment. Another complication that occurred in all diabetic rats, even in those that received polyphenols treatment, was diabetic cataract. Microscopic investigations reveal a crystalline cortex full of vacuoles or a homogeneous substance with residues of pathological crystalline fibres. Diabetic animals that benefited from polyphenols protection experience only scarce and isolated soft exudates.
The water-soluble crude polysaccharide GM, obtained from the Gynostemma pentaphyllum Makino by boiling-water extraction and ethanol precipitation, was fractionated by DEAE–Sepharose CL-6B column chromatography, and purified by Sephadex G-100 column chromatography, giving three polysaccharide fractions termed GMA, GMB and GMC. The monosaccharide components of them were studied by PC and GC. On the basis of superoxide radical assay, hydroxyl radical assay and self-oxidation of 1,2,3-phentriol assay, the antioxidant activities of GM, GMA, GMB and GMC were investigated. Among these contents, GMC had the higher scavenging effects on superoxide radicals and inhibitory effects on self-oxidation of 1,2,3-phentriol, and so should be explored as a novel potential antioxidant.