Phlomis armeniaca: Phenolic compounds, enzyme inhibitory and antioxidant activities

Industrial Crops and Products 78 (2015) 95–101

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Phlomis armeniaca: Phenolic compounds, enzyme inhibitory and antioxidant activities Cengiz Sarikurkcu a , Mehmet Cemil Uren b , Bektas Tepe c,∗ , Mustafa Cengiz d , Mehmet Sefa Kocak b a

Süleyman Demirel University, Faculty of Pharmacy, Department of Analytical Chemistry, Isparta, Turkey Süleyman Demirel University, Atabey Vocational School, Department of Medicinal and Aromatic Plants, Isparta, Turkey c Kilis 7 Aralık University, Faculty of Science and Literature, Department of Molecular Biology and Genetics, Kilis, Turkey d Süleyman Demirel University, Faculty of Science and Literature, Department of Chemistry, Isparta, Turkey b

a r t i c l e

i n f o

Article history: Received 30 July 2015 Received in revised form 29 August 2015 Accepted 8 October 2015 Keywords: Phlomis armeniaca Antioxidant Enzyme inhibitory Phenolic content HPLC

a b s t r a c t This study investigated the phenolic content, enzyme-inhibitory and antioxidant activities of ethyl acetate, methanol and water extracts of Phlomis armeniaca. HPLC analysis showed chlorogenic acid to be the most abundant phytochemical, followed by benzoic acid. Phosphomolybdenum and ␤-carotene bleaching tests indicated the ethyl acetate extract to have the best activity potential (1.80 mmol trolox equivalents TEs/g extract and 91.50%, respectively); however, the water extract performed best in 1,1diphenyl-2-picrylhydrazyl (DPPH) and superoxide anion radical scavenging tests (125.23 and 43.05 mg TEs/g extract, respectively). Chelating effects and reducing powers of the extracts were also determined. Ethyl acetate extract exhibited considerable enzyme inhibition potential on acetylcholinesterase, butyrylcholinesterase, tyrosinase, ␣-glucosidase and ␣-amylase (2.065 mg galantamine equivalents GALAEs/g extract, 4.579 mg GALAEs/g extract, 15.97 mg kojic acid equivalents/g extract, 0.474 mmol acarbose equivalents ACEs/g extract and 2.261 mmol ACEs/g extract, respectively). These findings suggest that P. armeniaca may be useful in the development of an alternative agent for oxidative stress, Alzheimer’s disease and diabetes. © 2015 Elsevier B.V. All rights reserved.

1. Introduction Phenolic compounds play a role in the biological activity of commonly consumed plants and are among the most important plant secondary metabolites (Alasalvar et al., 2001). The positive correlation between the antioxidant activities of plant species and the presence of phenolic compounds makes the identification of these molecules particularly important in terms of understanding the antioxidant potential of plant-based nutrients (Chan et al., 2011). In addition to their cardioprotective, neuroprotective and anti-carcinogenic activities, phenolic compounds are also natural preservative agents that prevent oxidative deterioration and microbial contamination of food (Crozier et al., 2009; Freeman et al., 2010). The Phlomis genus of the Lamiaceae family is a large genus with more than 100 known species distributed mainly throughout Eurasia and North Africa. Several Phlomis species such as

∗ Corresponding author. E-mail address: [email protected] (B. Tepe). http://dx.doi.org/10.1016/j.indcrop.2015.10.016 0926-6690/© 2015 Elsevier B.V. All rights reserved.

Pantoea stewartii and Pyrus syriaca are consumed in the form of herbal teas as remedies for gastrointestinal problems and as prophylactics against liver, kidney, bone and cardiovascular diseases (Carmona et al., 2005; Jabeen et al., 2013). In addition to their ethnopharmacological use, some species of Phlomis have demonstrated proven antidiabetic (Sarkhail et al., 2007), antinociceptive, antiulcerogenic, anti-inflammatory, antiallergenic (Sarkhail et al., 2003), anticarcinogenic (Kirmizibekmez et al., 2004), antioxidant and antimicrobial (Morteza-Semnani et al., 2004) activities. This study investigated three different Phlomis armeniaca extracts (ethyl acetate, methanol, and water) for phenolic contents [protocatechuic acid, (+)-catechin, p-hydroxybenzoic acid, chlorogenic acid, caffeic acid, (−)-epicatechin, ferulic acid, benzoic acid, rutin, rosmarinic acid, apigenin], enzyme inhibition (on acetyl cholinesterase, butyryl cholinesterase, tyrosinase, ␣-amylase and ␣-glucosidase), antioxidant activity (using ␤-carotene bleaching and phosphomolybdenum), radical scavenging (on DPPH, ABTS, superoxide-anion and nitric-oxide radicals), reducing power (by CUPRAC and FRAP assays) and chelating activities. Total phenolic contents as well as flavonoid, flavonol and saponin contents of the extracts were also identified. In addition, these findings were

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compared with reports in the literature on P. armeniaca phytochemical contents (Demirci et al., 2009; Saracoglu et al., 1995), enzyme inhibitory activity on ␣-amylase and ␣-glucosidase, and antioxidant activity (Dalar and Konczak, 2013). 2. Materials and methods 2.1. Plant material The aerial parts of P. armeniaca WILLD were randomly collected from Kemerkaya town, Bolvadin, Afyonkarahisar—Turkey, at the flowering stage, on 23 June 2013 (1520 m, 38◦ 53 59 N 31◦ 06 13 E). Taxonomic identification of the plant material was confirmed by the senior taxonomist Dr. Olcay CEYLAN, in Department of Biology, Mugla University, Mugla—Turkey. The voucher specimen was deposited at the Herbarium of the Department of Biology, Mugla University, Mugla—Turkey (Voucher No.: MUH1877). 2.2. Preparation of the solvent extracts To produce solvent extracts the air-dried samples (20 g) of the aerial parts of P. armeniaca were extracted by using a Soxhlet extractor for 5 h with 250 ml of solvents (ethyl acetate and methanol). For water extract, the air-dried samples (20 g) were extracted by boiling deionized water (200 ml) for 15 min. Ethyl acetate and methanol were then removed by a rotary evaporator. The water extract was freeze-dried. All extracts were stored at +4 ◦ C until analyzed. Yields of the solvent extracts from P. armeniaca are given in Table 1. 2.3. Total antioxidant activity Total antioxidant activities of the samples were evaluated by phosphomolybdenum (Zengin et al., 2014) and ␤-carotene bleaching (Sarikurkcu et al., 2012) methods. 2.4. Radical scavenging activity The effects of the samples on 1,1-diphenyl-2-picrylhydrazyl (DPPH) (Sarikurkcu, 2011), ABTS [2,2 Azino-bis (3ethylbenzothiazloine-6-sulfonic acid)] (Re et al., 1999), nitric oxide (• NO) reaction (Srivastava and Shivanandappa, 2011) and superoxide anion (O2 •− ) (Dasgupta and De, 2004) radicals were estimated according to the procedure in literature.

2.8. Determination of total bioactive components Total phenolic, flavonoid and saponin contents were determined by employing the methods given in the literature (Zengin et al., 2014). Total flavanol content was determined by employing the method given in the literature (Quettier-Deleu et al., 2000) with slight modification. Sample solution (0.25 ml) was added to 5 ml of 0.1% DMACA (p-dimethylaminocinnamaldehyde) in methanolic HCl reagent (3:1). The sample absorbance was read at 640 nm after 10 min incubation at room temperature. Total flavanol content was expressed as milligrams of (+)-catechin equivalents (mg CEs/g extract). 2.9. Quantification of phenolic compounds by RP-HPLC Phenolic compounds were evaluated by RP-HPLC (Shimadzu Scientific Instruments, Tokyo, Japan). Detection and quantification were carried out with a LC-10ADvp pump, a Diode Array Detector, a CTO-10Avp column heater, SCL-10Avp system controller, DGU-14A degasser and SIL-10ADvp auto sampler (Shimadzu Scientific Instruments, Columbia, MD). Separations were conducted at 30 ◦ C on Agilent® Eclipse XDB C-18 reversedphase column (250 mm × 4.6 mm length, 5 ␮m particle size). Phenolic compositions of the extracts were determined by a modified method of Caponio et al. (1999). Protocatechuic acid, (+)-catechin, p-hydroxybenzoic acid, chlorogenic acid, caffeic acid, (−)-epicatechin, ferulic acid, benzoic acid, rutin, rosmarinic acid and apigenin were used as standard (Fig. 1). Identification and quantitative analysis were done by comparison with standards (Figs. 2–4 ). The amount of each phenolic compound was expressed as milligram per gram of extract using external calibration curves, which were obtained for each phenolic standard. 2.10. Statistical analysis For all the experiments, all the assays were carried out in triplicate. The results are expressed as mean values and standard deviation (mean ± SD). The differences between the extracts were analyzed using one-way analysis of variance (ANOVA) followed by Tukey’s honestly significant difference post hoc test with ˛ = 0.05. This treatment was carried out using SPSS v. 14.0 program. 3. Results and discussion 3.1. Antioxidant activity

2.5. Reducing power The reducing power was investigated using cupric ion reducing (CUPRAC) (Apak et al., 2006) and ferric reducing antioxidant power (FRAP) (Aktumsek et al., 2013) methods, as previously described in the literature. 2.6. Metal chelating activity on ferrous ions Metal chelating activity on ferrous ions was evaluated by the method described by Aktumsek et al. (2013). 2.7. Enzyme inhibitory activity Enzyme inhibitory activities the samples were determined using cholinesterase (ChE), ␣-amylase, ␣-glucosidase (Zengin et al., 2014) and tyrosinase (Orhan et al., 2012) enzymes by employing the methods given in the literature.

Table 1 presents yield values obtained at the end of extraction as well as potential antioxidant activity of the extracts identified using phosphomolybdenum and ␤-carotene bleaching tests. The maximum extract yield was obtained with methanol (15.50% w/w), followed by water (14.05% w/w) and ethyl acetate extract (2.45% w/w). Phosphomolybdenum testing indicated ethyl acetate extract to exhibit a higher level of total antioxidant activity (1.80 ± 0.13 mmol TEs/g extract) than methanol and water extracts, which exhibited similar levels of total antioxidant activity (1.39 and 1.42 mmol TEs/g extract, respectively). Similarly, ␤-carotene bleaching tests indicated ethyl acetate extract to show the greatest total antioxidant activity (91.50%), followed by methanol (75.63%) and water extracts (50.30%). Total antioxidant activity of the ethyl acetate extract was significantly higher than that of both the methanol and water extracts (p < 0.05), and total antioxidant activity of the methanol extract was significantly higher than that of the water extract (p < 0.05). The reliability of data related to antioxidant activity was checked using BHA, BHT and Trolox as positive

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Table 1 Extraction yield and total antioxidant activity (by ␤-carotene bleaching and phosphomolybdenum methods) of P. armeniaca solvent extracts (mean ± SD).* Samples

Extraction yield (%)

Ethyl acetate Methanol Water BHA BHT Trolox

2.45 15.50 14.05 – – –

* ** ***

Total antioxidant activity Phosphomolybdenum(mmol TEs/g extract)**

␤-Carotene bleaching(%)***

1.80 ± 0.13a 1.39 ± 0.07b 1.42 ± 0.05b – – –

91.50 ± 1.82a,b 75.63 ± 0.51c 50.30 ± 2.90d 95.76 ± 1.26a 93.24 ± 0.46a,b 88.82 ± 1.78b

Different superscript letters in the same column indicate significant difference (p < 0.05). TEs, trolox equivalents. At 2 mg/ml concentration.

Fig. 1. Chromatographic profile of the chemical standards.

Fig. 2. Chromatogram of the ethyl acetate extract of P. armeniaca.

controls; accordingly, BHA and BHT showed 90% protection against linoleic acid oxidation. Table 2 shows the radical scavenging potential of the extracts in the presence of DPPH, superoxide anions, nitric oxide radicals and

ABTS cations. Whereas the water extract exhibited the best radical scavenging potential in DPPH (125.23 mg TEs/g extract) and superoxide anion radical scavenging (43.05 mg TEs/g extract) tests, the methanol extract performed best in ABTS cation (146.81 mg TEs/g

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Fig. 3. Chromatogram of the methanol extract of P. armeniaca.

Fig. 4. Chromatogram of the water extract of P. armeniaca. Table 2 Radical scavenging activity of P. armeniaca solvent extracts (mean ± SD).* Samples

DPPH radical (mg TEs/g extract)**

ABTS radical cation (mg TEs/g extract)**

Superoxide anion radical (mg TEs/g extract)**

Nitric oxide radical (mmol TEs/g extract)**

Ethyl acetate Methanol Water

35.98 ± 1.27c 67.48 ± 3.85b 125.23 ± 1.08a

100.48 ± 0.30b 146.81 ± 0.17a 97.98 ± 3.50b

15.73 ± 0.41c 37.64 ± 0.55b 43.05 ± 1.09a

na*** 1.59 ± 0.07a 0.70 ± 0.14b

* ** ***

Different superscript letters in the same column indicate significant difference (p < 0.05). TEs, trolox equivalents. na, not active.

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Table 3 Reducing power and metal chelating activities of P. armeniaca solvent extracts (mean ± SD).* Samples

CUPRAC(mg TEs/g extract)**

FRAP(mg TEs/g extract)**

Chelating effect(mg EDTAEs/g extract)***

Ethyl acetate Methanol Water

92.40 ± 6.78 127.60 ± 5.86b 127.77 ± 3.91b

59.40 ± 3.79 87.08 ± 1.92b 96.58 ± 3.79a

6.37 ± 1.21b 20.40 ± 1.00a 22.64 ± 0.65a

* ** ***

a

c

Different superscript letters in the same column indicate significant difference (p < 0.05). TEs, trolox equivalents. EDTAEs, disodium edetate equivalents.

Table 4 Enzyme inhibitory activity of P. armeniaca solvent extracts (mean ± SD).* Samples

Acetyl cholinesterase (mg GALAEs/g extract)**

Butyryl cholinesterase (mg GALAEs/g extract)**

Tyrosinase (mg KAEs/g extract)***

␣-Amylase (mmol ACEs/g extract)****

␣-Glucosidase (mmol ACEs/g extract)****

Ethyl acetate Methanol Water

2.065 ± 0.019a 1.585 ± 0.010b 0.534 ± 0.010c

4.579 ± 0.058a 1.436 ± 0.058b na*****

15.97 ± 0.94a na 9.88 ± 0.27b

0.474 ± 0.007a 0.335 ± 0.007b 0.069 ± 0.004c

2.261 ± 0.243b 6.055 ± 0.810a 3.339 ± 0.020a,b

* ** *** **** *****

Different superscript letters in the same column indicate significant difference (p < 0.05). GALAEs, galanthamine equivalents. KAEs, kojic acid equivalents. ACEs, acarboseequivalents. Na, not active.

extract) and nitric oxide radical tests (1.59 mmol TEs/g extract). Ethyl acetate extract showed no nitric oxide radical scavenging activity. Whereas DPPH, superoxide anion and nitric oxide radical scavenging tests indicated significant differences among all 3 extracts, no differences were found between the ethyl acetate and water extracts in terms of ABTS cation-scavenging potential (p < 0.05). Table 3 presents the results of cupric reducing antioxidant capacity (CUPRAC) and ferric reducing antioxidant power (FRAP) tests evaluating the reducing power of the P. armeniaca extracts as well as the metal chelating effects of the extracts. As the table shows, the water extract performed best in all three tests, followed by the methanol extract and the ethyl acetate extract. CUPRAC testing found the water extract to perform only slightly better (127.77 mg TEs/g extract) than the methanol extract (127.60 mg TEs/g extract), both of which performed significantly better than the ethyl acetate extract (92.40 mg TEs/g extract) (p < 0.05). FRAP testing showed the water extract (96.58 mg TEs/g extract) to have reducing power significantly greater than both the methanol extract (87.08 mg TEs/g extract) and the ethyl acetate extract (96.58 mg TEs/g extract) (p < 0.05). Results of metal chelating tests for water, methanol and ethyl acetate extracts were, respectively, 22.64, 20.40 and 6.37 mg EDTAEs/g extract. Whereas the difference in metal chelating ability of the water and methanol extracts showed no statistically significant difference, the differences between the water and ethyl acetate extracts and between the methanol and ethyl acetate extracts were statistically significant (p < 0.05). Dalar and Konczak (2013) reported the FRAP and oxygen radical scavenging capacities of P. armeniaca to be 850 ␮mol Fe2+ Eq./g dry weight and 3000 ␮mol Trolox Eq./g dry weight, respectively. However, given the differences in methodology used in identifying and presenting antioxidant activity, it is difficult to make quantitative comparisons between this and the preset study.

3.2. Enzyme inhibition capacity Type 2-diabetes can be managed by lowering blood sugar levels through the inhibition of enzymes involved in regulating carbohydrate metabolism and glucose release from food to suppress postprandial hyperglycemia. The main enzymes targeted by such treatment are ␣-glucosidase and ␣-amylase, which break down

large polysaccharide molecules into sugars (Sakulnarmrat and Konczak, 2012). Natural enzyme inhibitors can be used in the treatment of diabetes to effectively limit the release of glucose from starch (Apostolidis et al., 2006; Dalar and Konczak, 2013). Acetylcholine plays a role in cognitive functions, especially memory. Acetylcholinesterase (AChE) terminates acetylcholine action in cholinergic synapses. Cognitive impairment resulting from low levels of cholinergic neurotransmission is strictly associated with Alzheimer’s disease (Giacobini, 2003; Holzgrabe et al., 2007), which is characterized by decreases in acetylcholine levels due to the degeneration of cholinergic neurons in the basal forebrain. AChE inhibitors, which enhance cholinergic transmission by increasing synaptic acetylcholine levels, are frequently used in treating Alzheimer’s symptoms (Holzgrabe et al., 2007; Nordberg, 2006). The most common AChE inhibitors used in such treatment are galanthamine, donepezil and rivastigmine (Petzer et al., 2012). Moreover, inhibition of butyrylcholinesterase, another enzyme with an important role in Alzheimer’s disease, has been shown to reduce beta-amyloid protein levels and improve learning performance in rats (Greig et al., 2005). Tyrosinase is a bifunctional, copper-containing enzyme found in numerous plants, bacteria, fungi, animals and insects. In the presence of oxygen, tyrosinase catalyses two reactions: hydroxylation of monophenols to produce diphenols, followed by the oxidation of o-diphenols to produce quinines (Garcia-Molina et al., 2005). Contact between quinine and organic molecules such as amines, amino acids, peptides and proteins produces a reaction that initiates deterioration in foods, including browning and loss of nutritional quality (Carpenter, 1981). Table 4 shows the inhibitory activities of P. armeniaca ethyl acetate, methanol and water extracts on acetylcholinesterase, butyrylcholinesterase, tyrosine, ␣-amylase and ␣-glucosidase. The ethyl acetate extract exhibited the strongest inhibitory activity on all enzymes tested with the exception of ␣-glucosidase. The ethyl acetate extract showed the greatest activity against tyrosinase (15.97 mg KAEs/g extract), followed by butyrylcholinesterase (4.579 mg GALAEs/g extract), acetylcholinesterase (2.065 mg GALAEs/g extract) and ␣-amylase (0.474 mmol ACEs/g extract). With regard to ␣-glucosidase inhibition, the methanol extract exhibited the strongest inhibitory activity against ␣glucosidase (6.055 mmol ACEs/g extract), followed by the water

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Table 5 Total bioactive components of P. armeniaca solvent extracts (mean ± SD).* Samples

Total phenolic(mg GAEs/g extract)**

Total flavonoid(mg REs/g extract)***

Total flavanol(mg CEs/g extract)****

Total saponin(mg QAEs/g extract)*****

Ethyl acetate Methanol Water

41.99 ± 2.14 55.22 ± 1.95a 54.39 ± 2.77a

29.04 ± 4.30 53.74 ± 1.66a 32.14 ± 1.13b

3.58 ± 0.19 1.20 ± 0.05b 0.23 ± 0.04c

334.26 ± 21.39a 295.37 ± 47.58a 160.19 ± 3.93b

* ** *** **** *****

b

b

a

Different superscript letters in the same column indicate significant difference (p < 0.05). GAEs, gallic acid equivalents. REs, rutin equivalents. CEs, catechin equivalents. QAEs, quillaja equivalents.

Table 6 Phenolic components in P. armeniaca solvent extracts (mean ± SD)* and analytical characteristics for determination of phenolics. No.

1 2 3 4 5 6 7 8 9 10 11 * **

Phenolic components

Protocatechuic acid (+)-Catechin p-Hydroxybenzoic acid Chlorogenic acid Caffeic acid (−)-Epicatechin Ferulic acid Benzoic acid Rutin Rosmarinic acid Apigenin

Concentration (mg/g extract)

Analytical characteristics

Ethyl acetate

Methanol

0.37 ± 0.022 2.00 ± 0.070a 0.57 ± 0.020a 3.21 ± 0.060c 0.43 ± 0.010a 0.13 ± 0.007a nd 0.65 ± 0.020b 0.98 ± 0.040a nd 0.59 ± 0.010a

0.26 ± 0.005 0.77 ± 0.016c nd** 11.95 ± 0.919a 0.15 ± 0.011b nd 0.29 ± 0.011a nd 0.85 ± 0.016b nd nd

a

Water b

0.28 ± 0.005 0.98 ± 0.018b 0.16 ± 0.001b 8.58 ± 0.125b 0.39 ± 0.012a nd 0.21 ± 0.012a 6.14 ± 0.089a 1.09 ± 0.018a nd nd b

Linear range(mg/l)

Linear equation

R2

LOD(mg/l)

LOQ(mg/l)

0.20–25.0 0.90–113 0.20–25.0 0.35–45.0 0.16–21.0 0.50–66.0 0.12–17.0 0.85–55.0 0.40–55.0 0.40–55.1 0.17–11.0

y = 48,107x − 11,153 y = 20,346x − 29,275 y = 62,896x − 11,801 y = 37,172x − 20,503 y = 101,382x − 18,712 y = 30,982x − 22,609 y = 94,621x − 15,153 y = 9578.2x − 2819.6 y = 26,877x − 18,033 y = 39,556x − 27,953 y = 95,601x − 6571.1

0.9991 0.9988 0.9994 0.9988 0.9993 0.9990 0.9993 0.9998 0.9990 0.9989 0.9997

0.086 0.172 0.007 0.080 0.054 0.170 0.004 0.111 0.180 0.050 0.034

0.260 0.522 0.020 0.241 0.162 0.514 0.011 0.335 0.550 0.150 0.104

Different superscript letters in the same column indicate significant difference (p < 0.05). nd, not detectable.

extract (3.339 mmol ACEs/g extract) and the ethyl acetate extract (2.261 mmol ACEs/g extract). Furthermore, as Table 4 shows, the water extract exhibited no activity against butyrylcholinesterase, and the methanol extract showed no activity against tyrosinase. With the exception of activity against ␣-glucosidase, differences in inhibitory activity among extracts were statistically significant (p < 0.05). Dalar and Konczak (2013) reported that a hydrophilic lyophilized extract of P. armeniaca exhibited weak inhibitory activity against ␣-amylase and pronounced inhibitory activity against ␣-glucosidase, which suggests potential anti-diabetic properties. 3.3. Phenolic composition Table 5 presents data on the phenolic, flavonoid, flavonol and saponin contents of the 3 extracts tested. The methanol extract was found to contain larger amounts of phenolic (55.22 mg GAEs/g extract) and flavonoid (53.74 mg REs/g extract) compounds than the water and ethyl acetate extracts, whereas the ethyl acetate extract had higher flavonol (3.58 mg CEs/g extract) and saponin (334.26 mg QAEs/g extract) contents. A linear relationship is known to exist between a plant’s phytochemical contents and its antioxidant activity. While the literature contains numerous studies demonstrating the antioxidant potential of flavonoids and related compounds as well as phenolic acids, relatively few studies have investigated the potential of saponins. The amphipathic nature of saponins allows them to act as surfactants to enhance penetration of various compounds through cell membranes (Sigma–Aldrich, 2009). For this reason, saponins are assumed to facilitate the entrance of biologically active compounds into cells, thereby increasing the intensity of action. While qualitative spectrophotometric data is usually insufficient for establishing a clear relationship between phytochemical content and biological activity, in many cases, HPLC can determine the phytochemical profile of plant samples. However, the number and types of components identified strictly depends on the standards used and the richness of the library of instruments. Quantitative chromatographic techniques such as flash column chromatography and

bioactivity-guided fractionation are the most practical and consistent means for identifying the specific phytochemical components responsible for a particular biological activity. In this study, the phenolic acids, flavonoids and related compounds of P. armeniaca were measured quantitatively (Table 6). The most abundant phytochemical identified was chlorogenic acid, followed by benzoic acid. Of the 3 extracts tested, ethyl acetate extract contained the highest amounts of protocatechuic acid (0.37 mg/g extract), (+)-catechin (2.00 mg/g extract), p-hydroxybenzoic acid (0.57 mg/g extract), caffeic acid (0.43 mg/g extract), (−)-epicatechin (0.13 mg/g extract) and apigenin (0.59 mg/g extract), whereas the methanol extract contained the highest amounts of chlorogenic acid (11.95 mg/g extract) and ferulic acid (0.29 mg/g extract), and the water extract contained the highest amounts of benzoic acid (6.14 mg/g extract) and rutin (1.09 mg/g extract). A previous study conducted by Demirci et al. (2009) using 1D and 2D NMR, FT-IR, UV and HRMS reported two distinct phytochemicals available in P. armeniaca, namely (−)-8(14), 15-isopimaradien-11 alpha-ol and 4-Methoxycarbonyl-7-methyl cyclopenta[c]pyrane-a fulvoiridoid. In contrast, Saracoglu et al. (1995) isolated the following 10 known glycosidic compounds from methanolic extracts of P. armeniaca: betulalbuside A;8hydroxylinaloyl,3-o-beta-d-glucopyranoside; Ipolamide; Acteoside; leucosceptoside A; martynoside; forsythoside B; phlinoside B; phlinoside C; teuerioside. A literature review uncovered no other studies on this subject, so the present study is assumed to be the first to report on the phenolic compounds of P. armeniaca. 4. Conclusions This study examined the phytochemical properties of P. armeniaca extractions in ethyl acetate, methanol and water. The P. armeniaca ethyl acetate extract was found to exhibit the most activity, including considerable inhibitory activity on the enzymes tested. According to the results, P. armeniaca comprises a rich mixture of phytochemicals that offer comprehensive protection

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