Home | About PR | Editorial board | Search | Ahead of print | Current Issue | Archives | Instructions | Subscribe | Advertise | Contact us |   Login 
Pharmacognosy Magazine
Search Article 
  
Advanced search 
 


 
 Table of Contents 
ORIGINAL ARTICLE
Year : 2018  |  Volume : 10  |  Issue : 4  |  Page : 368-378  

HPLC-DAD-ESI-MS/MS characterization of bioactive secondary metabolites from Strelitzia nicolai leaf extracts and their antioxidant and anticancer activities In vitro


1 Department of Medicinal Chemistry, Theodor Bilharz Research Institute, Giza, Egypt
2 Department of Medicinal Chemistry, Theodor Bilharz Research Institute, Giza, Egypt; Department of Chemistry, College of Science and Arts, Sajir, Shaqra University, Shaqra, Kingdom of Saudi Arabia
3 Department of Biochemistry and Molecular Biology, Theodor Bilharz Research Institute, Giza, Egypt

Date of Web Publication26-Oct-2018

Correspondence Address:
Dr. Mosad Ghareeb
Department of Medicinal Chemistry, Theodor Bilharz Research Institute, Kornish El-Nile Street, Warrak El-Hader, Imbaba, P. O. 12615, Giza
Egypt
Login to access the Email id

Source of Support: None, Conflict of Interest: None


DOI: 10.4103/pr.pr_89_18

Rights and Permissions
   Abstract 


Background: Strelitzia nicolai Regel and Körn (Strelitziaceae) is native to Southern Africa whose phytochemistry and pharmacology were slightly investigated. Materials and Methods: In the current work, different solvent extracts of S. nicolai were screened for their chemical profiles through high-performance liquid chromatography coupled with diode array detection and electrospray ionization mass spectrometry (HPLC-DAD-ESI-MS/MS) analyses. Furthermore, their in vitro antioxidant, cytotoxic, and anticancer activities were evaluated using 2,2'-diphenyl-1-picrylhydrazyl radical (DPPH), 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) & ferric reducing antioxidant power (FRAP) and crystal violet staining (CVS) colorimetric assays, respectively. Results: HPLC-DAD-ESI-MS/MS analyses led to the identification of nineteen and eleven phenolic compounds from the ethyl acetate and n-butanol extracts, respectively including flavonoids (e.g., quercetin 3-(2 G-rhamnosylrutinoside, quercetin, quercetin-3-O-glucoside, kaempferol-3,7-O-dirhamnoside, isorhamnetin-3-O-rutinoside and kaempferol-3-O-glucoside), phenolic acids derivatives (e.g., chlorogenic acid glycoside, protocatechuic acid-O-glucoside and caftaric acid), chalcones (e.g., xanthoangelol), and phenylethanoids (e.g., ligstroside glucoside). Moreover, in the DPPH assay the IC50value of the most active ethyl acetate extract was 20.49 μg/mL, relative to 2.92 μg/mL of ascorbic acid. ABTS and FRAP results reinforced the results of DPPH assay. According to the National Cancer Institute criteria, the tested extracts showed weak to moderate cytotoxic activities with IC50values ranged from 65.23 to 451.29 μg/mL. Furthermore, the EtOAc and n-BuOH extracts showed a noticeable anticancer activity with CVS spectroscopic readings for liver hepatocellular carcinoma growth 0.806 and 0.684 at a concentration (125 μg/mL), as well as 0.730 and 0.618 at concentration (500 μg/mL), respectively against control at 1.022. Conclusion: The obtained results reveal the high efficacy of the phenolic-rich extracts from S. nicolai as naturally occurring antioxidant and anti-tumor agents.
Abbreviations Used: HPLC-DAD-ESI-MS/MS: High-performance liquid chromatography-diode array detection-electrospray ionization-mass/mass; DPPH: 2,2'-Diphenyl-1-picrylhydrazyl radical; ABTS: 2,2'-Azino-bis (3-ethylbenzothiazoline-6-sulphonic acid); FRAP: Ferric reducing antioxidant power; TPTZ: Tripyridyl-s-triazine; FE: Ferrous equivalents; DMEM: Dulbecco's modified eagle's medium; DMSO: Dimethyl sulfoxide; EDTA: Ethylenediaminetetraacetic acid; PBS: Phosphate buffered saline; HepG-2: Liver hepatocellular carcinoma; CVS: Crystal violet stain; NCI: National cancer institute; Glu: Glucose; Rha: Rhamnose.

Keywords: Anticancer activity, antioxidant activity, cytotoxicity, HPLC-DAD-ESI-MS/MS, polyphenolics, Strelitzia nicolai


How to cite this article:
Ghareeb M, Saad A, Ahmed W, Refahy L, Nasr S. HPLC-DAD-ESI-MS/MS characterization of bioactive secondary metabolites from Strelitzia nicolai leaf extracts and their antioxidant and anticancer activities In vitro. Phcog Res 2018;10:368-78

How to cite this URL:
Ghareeb M, Saad A, Ahmed W, Refahy L, Nasr S. HPLC-DAD-ESI-MS/MS characterization of bioactive secondary metabolites from Strelitzia nicolai leaf extracts and their antioxidant and anticancer activities In vitro. Phcog Res [serial online] 2018 [cited 2018 Dec 16];10:368-78. Available from: http://www.phcogres.com/text.asp?2018/10/4/368/244099





SUMMARY

  • The current research work evaluated the biological activities of different solvent extracts of Strelitzia nicolai including antioxidant, anticancer, and cytotoxic activities
  • Among the tested extracts, the ethyl acetate and n-butanol extracts are the most promising extracts
  • High-performance liquid chromatography coupled with diode array detection and electrospray ionization mass spectrometry analyses of the most active extracts led to the characterization of certain polyphenolic compounds, the majority are flavonoids and phenolic acids.



   Introduction Top


Strelitziaceae is a tropical monocotyledonous ornamental family famous by its bioactive compounds namely phenalenones.[1]Strelitzia nicolai Regel and Körn are usually known as the white bird of paradise tree.[2] This plant is native to Southern Africa and broadly growing in many tropical regions around the world.[3] To the best of our knowledge, there is limited information available in the literature about the phytochemical and biological investigations were reported on the plant. Bilirubin-IX is orange pigment with cyclic tetrapyrrole nucleus was isolated from the arils of S. nicolai.[4],[5],[6],[7] Moreover, it was reported that this pigment showed potent antioxidant and anticancer activities.[7] In addition, the main chemical ingredients of the essential oil isolated from the seed arils of the plant were categorized as follows: amine, ethers, ketones, hydrocarbons, aromatic compounds, alcohols, amides, and esters.[3] Moreover, accumulation of the reactive species in our bodies led to oxidative stress, which is associated with several disorders like cancer and cardiovascular diseases.[8] Cancer is a well-known global problem which represents the second cause of death and accounts for “7–8 million deaths” worldwide.[9] The treatment of cancer is based on the use of synthetized chemotherapeutic drugs, using this drugs as chemopreventive agents accompanied by a series of health problems due to their side effects and low safety, this concept encourage scientists to discover the role of medicinal plants in cancer therapeutic as natural sources of naturally occurring anticancer agents.[9],[10] Recently, there is increasing interest in the chemical investigation and characterization of the different class of secondary metabolites especially polyphenolic compounds to establish their structure-activity relationship.[11] Since there no adequate information has been documented on the chemical and biological profile of the plant, therefore, the aim of the current study is to characterize the polyphenolic compounds of the different solvent extracts from S. nicolai leaves via high-performance liquid chromatography coupled with diode array detection and electrospray ionization mass spectrometry (HPLC-DAD-ESI-MS/MS) analysis as well as their antioxidant and anticancer activities.


   Materials and Methods Top


Plant material

The fresh leaves S. nicolai were collected from Zoo Garden, Giza, Egypt in June 2014. The identity of the plant was established by Dr. Tearse Labib, Botany Specialist, Department of Flora and Taxonomy, El-Orman Botanical Garden, Giza, Egypt. A voucher specimen (No. S25/5/6) was kept at the herbarium of the garden.

Extraction and fractionation

Dry powdered leaves of S. nicolai (1.5 kg), were extracted with MeOH in room temperature with shaking day by day followed by filtration and again extraction for 4 times. The extract was filtered using Whatman filter paper No. 1 and concentrated using Rotatory evaporator (Buchi, Switzerland) at (40°C ± 2°C). The crude extract was collected and stored at room temperature in the dark for the further process. The methanolic crude extract (230 g) was defatted by washing several times with petroleum ether (60–80° C), then undergoing fractionation process using organic solvents such as CH2Cl2; EtOAc; and n-BuOH (5 mL × 150 mL solvent).

High-performance liquid chromatography coupled with diode array detection and electrospray ionization mass spectrometry conditions

The phytochemical analysis of polyphenolic compounds was done using HPLC-photodiode array (PDA)-MS/MS. The liquid chromatography system was Thermofingan (Thermo Electron Corporation, USA) coupled with an LCQ Duo ion trap MS with an ESI source (ThermoQuest). The separation was achieved by using a C18 reversed-phase column (Zorbax Eclipse XDB-C18, Rapid resolution, 4.6 mm × 150 mm, 3.5 μm, Agilent, USA). A gradient of water and acetonitrile (with 1% formic acid each in the positive mode) was applied from 5% to 50% ACN in 60 min with flow rate 1 mL/min throughout the whole run. The samples were injected automatically using autosampler surveyor ThermoQuest. The instrument was controlled by Xcalibur software (Thermo Fisher Scientific Inc., USA) to collect the ultraviolet (UV) chromatogram using PDA mode. The MS operated in the negative mode with a capillary voltage of − 10 V, a source temperature of 200°C, and high-purity nitrogen as a sheath and auxiliary gas at a flow rate of 80 and 40 (arbitrary units), respectively. The ions were detected in a full scan mode and mass range of 50–2000 m/z.

Antioxidant assays

2,2'-Diphenyl-1-picrylhydrazyl radical free radical-scavenging assay

2,2'-Diphenyl-1-picrylhydrazyl radical (DPPH) assay was performed according to the method described by Ghareeb et al. (2018). Briefly, 200 μl of plant extract, diluted appropriately in methanol in a concentration range from 0.24 to 500 μg/mL, was mixed with 100 μl of 0.2 mM DPPH in methanol in wells of 96-well plates. The plates were kept in the dark for 15 min; thereafter, the absorbance of the solution was measured at 515 nm in a Biochrom Asys UVM 340 Microplate Reader. Appropriate blanks, methanol, and standards (ascorbic acid solutions in methanol) were analyzed simultaneously. The scavenging activity (in %) was calculated using the following equation:

DPPH scavenging (%) = 100 × [(Abs sample + DPPH) - (Abs sample blank)]/[(Abs DPPH) - (Abs methanol)]

The IC50 value is defined as the amount of extract needed to scavenge 50% of DPPH radicals. All analyses were performed in triplicate.[12]

2,2'-Azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) assay

The samples were dissolved in water to prepare the stock solutions (1 mg/mL) from which a radical-scavenging activity was determined by the 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) (ABTS)+ radical cation decolorization assay,[13] over a concentration range of 0.24–500 μg/mL. The ABTS cation radical was prepared by reacting 7 mM aqueous solution of ABTS (15 mL) with 140 mM potassium persulfate (264 μl). The mixture was allowed to stand in dark at room temperature for 16 h before use. Before assay, the ABTS working reagent was diluted with methanol to give an absorbance of 0.70 ± 0.02 at 734 nm and was equilibrated at room temperature. The reaction mixtures in the 96-well plates consisted of sample (50 μl) and the ABTS methanol working solution (100 μl). The mixture was stirred and left to stand for 10 min in dark, and then the absorbance was determined at 734 nm against a blank. All determinations were performed in triplicate. The scavenging activity (in %) was calculated as follows:

% scavenging rate = [1− (A1–A2)/A0] ×100.

Where A0 is the absorbance of the control (without sample), and A1 is the absorbance in the presence of the sample, A2 is the absorbance of the sample without ABTS working solution. The scavenging activity of the samples was expressed as IC50 value, which is the effective concentration, at which 50% of ABTS radicals were scavenged. Trolox was used as a standard.

Ferric reducing antioxidant power assay

The ferric reducing antioxidant power (FRAP) assay was carried out according to the previously reported procedure,[14] with minor modifications. Each sample was dissolved in methanol to prepare the stock solution (1 mg/mL). Briefly, the working FRAP reagent was prepared freshly by mixing 300 mM acetate buffer (pH 3.6), a solution of 10 mM 2, 4, 6-tripyridyl-s-triazine (TPTZ) in 40 mM hydrochloric acid and 20 mM ferric chloride at 10:1:1 (v/v/v). 20 μl of each extract was mixed with 180 μl FRAP reagent in wells of 96-well plates. The mixture was then incubated for 6 min at 37°C, and the absorbance was measured at 595 nm in a microplate reader (Biochrom Asys UVM 340). Appropriate blanks of plant extract and FRAP reagent lacking TPTZ (to correct the colors of the extracts) were run, together with quercetin (in methanol), and ferrous sulfate heptahydrate (FeSO4.7H2O) was used as a standard. FRAP activity was calculated as ferrous equivalents, the concentration of extract/quercetin which produced an absorbance value equal to that of 1 mM FeSO4.

Evaluation of cytotoxic activities

Splenocytes were isolated from normal albino mouse according to Goodman et al.[15] beginning with washing thoroughly with 70% alcohol whole body and cervical dislocation done after anesthesia, and then the abdominal cavity was incised, and the spleen was transferred to sterile  Petri dish More Details and slice into small pieces.[16] Fragments were placed onto a strainer attached to a 50-mL conical tube. Excised spleen pieces were pressed through a strainer using a plunger end of a syringe and cells have been washed with phosphate buffered saline (PBS) (Dulbecco's PBS, pH = 7.4). Cell suspension centrifuged at 1600 rpm for 5 min. Cells pellets were resuspended in 2 mL lysing buffer (0.15 M NH4Cl, 1 mM KHCO3, and 0.1 mM ethylenediaminetetraacetic acid). The cells have been incubated in a 37°C water bath for 2 min, and 30 mL of PBS were added and cells centrifuged at 1600 rpm for 5 min. Pellet cells were resuspended in 3 mL growth Dulbecco's Modified Eagle's Medium, Lonza, Belgium), 30% FBS (Hyclone, USA), 1% Penicillin/Streptomycin (Lonza, Belgium), and 1% L-glutamine (Lonza, Belgium)[17],[18] at final concentration of 2000 × 10000 cells per mL, the cells were cultured in 96 well plates. Cell count was performed and viability checked using trypan blue and a hemacytometer. Cells were incubated with serial dilutions of eleven fractions dissolved in dimethyl sulfoxide (DMSO) as follow (1000 μg/mL, 500 μg/mL, 250 μg/mL, 125 μg/mL, 62.5 μg/mL, and 31.2 μg/mL) at 37°C in 5% CO2 and 90% humidified atmosphere for 36 h.

Evaluation of anticancer activities

Liver hepatocellular carcinoma (HepG-2) cell line obtained from the holding company for biological products and vaccines, Egypt (VACSERA) passage number 80 ≈ 85 has been cultured in a T25 flask with Roswell Park Memorial Institute medium contains 10% fetal bovine serum, 100 U/mL penicillin and 100 μg/mL streptomycin and incubated in a humidified 5% CO2 incubator at 37°C.[19] Culture medium was removed after reaching 90% confluency and 0.25% trypsin (Gibco/Invitrogen) was added and flask observed under an inverted microscope until all cells were detached. Cells were cultured in 96-well plates at a density of 10000 cells per well. Incubation has been done for 48 h until sheet was observed. Serial dilutions of fractions in DMSO were added to cultured cells at concentrations of 1000 μg/mL, 500 μg/mL, 250 μg/mL, 125 μg/mL, 62.5 μg/mL, and 31.2 μg/mL and incubated for 24 h. Media were decanted and plates were washed carefully with PBS (Dulbecco's PBS, pH = 7.4) and cells were stained with 20 μl of 0.5% crystal violet (Sigma-Aldrich Corp., St. Louis, MO, USA) in 30% ethanol for 10 min at room temperature. Plate wells were washed 3 times using distilled water[20] and the absorbance was measured at optical density = 490 nm.

Statistical analysis and dose-response curve

A dose-response curve has been traced to find the equation and estimate IC50 using unpaired Student's t-test excel, referring to different concentrations, more than three to draw the curve which presented as mean standard deviation (SD) taking in consideration P < 0.05.


   Results and Discussion Top


Antioxidant activity

Different solvent extracts of S. nicolai were evaluated for their antioxidant activities using three techniques including; DPPH, ABTS, and FRAP. In DPPH assay, the IC50 values for the tested extracts ranged from 20.49 to 118.17 μg/mL compared to ascorbic acid as standard with IC50 equal to 2.92 μg/mL. The results are in the order: EtOAc > n-BuOH> MeOH> H2O > CH2Cl2> Pet. ether. In addition, all tested extracts showed similar activity using ABTS assay with IC50 values arranged in the following order: EtOAc (9.18) > n-BuOH (12.43) > MeOH (15.73) > H2O (23.18) > CH2Cl2 (29.89) > Pet. ether extracts (52.25) (μg/mL) compared to Trolox (IC50 = 1.63 μg/mL). In FRAP assay, the EtOAc fraction showed high reducing power activity with 22.19 mM FeSO4 equivalent/mg extract, followed by n-BuOH (18.34), MeOH (15.46), H2O (11.39), CH2Cl2 (7.92), and Pet. ether extracts (1.58), respectively, compared to quercetin (21.45). Results are documented in [Table 1]. In conclusion, the results of the three assays agree with each other and the high antioxidant activity of the EtOAc and n-BuOH extracts may be due to the presence of the tentatively identified polyphenolic compounds which are broadly reported in the literature by their antioxidant potential for instance; flavonoids (e.g. quercetin 3-(2 G-rhamnosylrutinoside), rutin, quercetin-3-O-glucoside, kaempferol-3,7-O-dirhamnoside, isorhamnetin-3-O-rutinoside, kaempferol-3-O- rutinoside, and kaempferol-3-O-glucoside),[21],[22] and phenolic acids (e.g. chlorogenic acid glycoside, sinapaldehyde, and caftaric acid).[23],[24]
Table 1: Antioxidant activities of different extracts of S. nicolai

Click here to view


In vitro cytotoxic and anticancer activities of different solvent extracts

Different solvent extracts of S. nicolai have been evaluated for their in vitro cytotoxic potential to murine spleen cells by visual counting for the vital cells after growing under the effect of serial dilutions of these extracts. The results revealed that methanol (1) and n-butanol (5) extracts showed very rare cytotoxic activities with IC50 values of451.29 and 382.38μg/mL, respectively. While pet. ether (2), dichloromethane (3), and ethyl acetate (4) extracts showed moderate cytotoxic effects with IC50 values of68.03, 90.29, and 65.23 μg/mL, respectively [Table 2] and [Figure 1], indicating that flagging cytotoxic effects against murine spleen cell have been observed. Moreover, the National Cancer Institute instructions demonstrate the criteria and the conditions of cytotoxic activity for a chemical complexes as the concentration of an inhibitor that is required for 50% inhibition of cells growth.[25] IC50 where the values ≤ 20 μg/mL, is considered to be potentially cytotoxic, while IC50 values 21-100 μg/mL = moderately cytotoxic, IC50 101–200 μg/mL = weakly cytotoxic and IC50> 501 μg/mL = no cytotoxic effect.[26],[27] On the other hand, significant anticancer effects against HepG-2 cell line after 24 h exposure were obvious using colorimetric assay. The results revealed that the crystal violet stain spectroscopic readings for HepG-2 growth at complex concentration (125 μg/mL) were 0.883, 0.931, 0.753, 0.806, and 0.684, while at complex concentration (500 μg/mL) were 0.684, 0.611, 0.702, 0.730, and 0.618, respectively for MeOH, pet. ether, dichloromethane, ethyl acetate and n-butanol extracts against control at 1.022 [Table 3]. Results showed an intimate relationship between anticancer activity and concentration. In general, the anticancer activity of the polyphenolic-rich extracts may be due to the presence of such phenolic compounds.[28],[29],[30],[31],[32] Moreover, these polyphenolic compounds are capable of inhibiting cancer cells through several modes of actions.[26]
Table 2: Different solvent extracts of S. nicolai showing the variation in cytotoxic effect (IC50) on murine spleen cells by visual counting under the inverted microscope

Click here to view
Figure 1: Cytotoxic effect of different solvent extracts of S. nicolai on murine splenocytes after 36 h incubation

Click here to view
Table 3: Crystal violet staining and colorimetric assay at optical density= 490 nm for human hepatoma cell line under the effect of S. nicolai extracts showing cytotoxicity profile

Click here to view


Characterization of the phenolic compounds in Strelitzia nicolai

The ethyl acetate and n-butanol extracts of S. nicolai were investigated for their polyphenolic constituents using HPLC-DAD-ESI-MS/MS technique. Nineteen compounds were detected and tentatively identified in the ethyl acetate extract were categorized as phenolic acids, flavonoids (glycosides and aglycones), chalcones, and other nucleus. On the other hand, eleven compounds were detected and tentatively identified in the n-butanol extract were categorized as phenolic acids and flavonoids (glycosides and aglycones). The identification of the phenolic compounds was based on comparing their fragmentation pattern using negative ion ionization mode with the available data in the literature. The chemical structures of some selected compounds and their full MS/MSn pattern will be mentioned below [Table 4], [Table 5] and [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6], [Figure 7].
Table 4: Phenolic compounds tentatively identified in the EtOAc extract of S. nicolai leaves by HPLC-DAD-ESI-MS/MS in negative ion mode

Click here to view
Table 5: Phenolic compounds tentatively identified in the n-butanol extract of S. nicolai leaves by HPLC-DAD-ESI-MS/MS in negative ion mode

Click here to view
Figure 2: Negative HPLC-DAD-ESI-MS/MS profiles of phenolic compounds were detected in the EtOAc extract of S. nicolai leaves

Click here to view
Figure 3: Mass spectrometry spectra and postulated fragmentation pattern of some selected phenolic compounds detected in the EtOAc extract of S. nicolai leaves using HPLC-DAD-ESI-MS/MS in negative ionization mode; (a) quercetin [M-H] m/z = 301; (b) quercetin-3-O-glucoside [M-H] m/z = 463; (c) rutin [M-H] m/z = 609; (d) isohammetin-3-O-rutinoside [M-H] m/z = 623; (e) kaempferol-3-O-glucoside [M-H] m/z = 447; and (f) kaempferol-3-O-rutinoside [M-H] m/z = 593

Click here to view
Figure 4: A proposed fragmentation pattern of compound 5 (rutin) and compound 10 (Kaempferol-3-O-rutinoside)

Click here to view
Figure 5: Negative HPLC-DAD-ESI-MS/MS profiles of phenolic compounds were detected in the EtOAc extract of S. nicolai leaves

Click here to view
Figure 6: Mass spectrometry spectra and postulated fragmentation pattern of some selected phenolic compounds detected in the n-BuOH extract of S. nicolai leaves using HPLC-DAD-ESI-MS/MS in negative ionization mode; (a) protocatechuic acid-O-glucoside [M-H] m/z = 315; (b) quercetin 3-(2 G-rhamnosylrutinoside) [M-H] m/z = 755; (c) kaempferol 3-O-rutinoside-7-O-rhamnoside [M-H] m/z = 739, and (d) ligstroside glucoside [M-H] m/z = 685

Click here to view
Figure 7: Chemical structures of some identified phenolic compounds in EtOAc and n-BuOH extracts of S. nicolai leaves

Click here to view


Phenolic compounds were detected in the EtOAc extract

Phenolic acids and their derivatives

Compound 1 was detected at Rt = 1.43 min, it showed a deprotonated ion [M-H] at m/z 515, it also generated a fragment ion as a base peak at m/z 341 corresponding to the loss of quinic acid moiety (-m/z 174 u) [M-H-quinic acid moiety], further loss of glucosyl moiety (-m/z 162 u) was confirmed by the appearance of a fragment ion at m/z 179 [M-H-quinic acid-glucose moiety]. Therefore, compound 1 could be identified as chlorogenic acid glucoside as previously described.[33] Compound 16 was detected at Rt = 28.92 min, it showed a deprotonated ion as a base peak [M-H] at m/z 177, it also generated a fragment ion at m/z 162 corresponding to the loss of methyl moiety (-m/z 15) [M-H-CH3], further loss of hydroxyl moiety was confirmed by the appearance of a fragment ion at m/z 145 [M-H-CH3-OH], further neutral loss of CO moiety of the aldehyde group was confirmed by the appearance of a fragment ion at m/z 117 (-m/z 28 u) [M-H-CH3-OH-CO]. Therefore, compound 16 could be identified as 3-(4-hydroxy-3-methoxyphenyl) prop-2-enal (coniferyl aldehyde).[34]

Compound 17 was detected at Rt = 30.57 min, it showed a deprotonated ion [M-H] at m/z 207, it also generated a fragment ion as a base peak at m/z 192 corresponding to the presence of sinapoyl moiety and due to the loss of methyl moiety (-m/z 15 u) [M-H-CH3], further loss of methyl moiety (-m/z 15 u) was confirmed through the fragment ion at m/z 177 [M-H-2CH3]. Thus, compound 17 could be identified as 3,5-dimethoxy-4-hydroxycinnamaldehyde (sinapaldehyde).[35] Compound 19 was detected at Rt = 40.70 min, it showed a deprotonated molecular ion [M-H] at m/z 311, it also generated a fragment ion at m/z 179 corresponding to the loss of tartaric moiety (-m/z 132 u) [M-H-tartaric], in addition to a fragment ion was appeared at m/z 149 corresponding to the loss of caffeoyl moiety (-m/z 162 u) [M-H-caffeoyl]. This fragmentation pattern was typically assigned to caftaric acid (cis-caffeoyl tartaric acid).[35],[36]

Flavonoids

Compound 3 was detected at Rt = 14.66 min, it showed a deprotonated molecule [M-H] at m/z 755, a MS/MS fragment was observed at m/z 609 was assigned to rutin molecule after the neutral loss of one rhamnosyl moiety (-m/z 146 u) [M-H-rhamnose moiety], a fragment ion was detected at m/z 591 due to further loss of water molecule (-m/z 18 u) [M-H-rhamnose moiety-H2O], another fragment ion was also appeared at m/z 445 due to the loss of the second rhamnosyl moiety (-m/z 146 u) [M-H-2rhamnose moiety-H2O], a diagnostic fragment ion was observed at m/z 301 was assigned to quercetin a glycone and can be explained by the release of the glucose moiety (-m/z 162 u) [M-H-2rhamnose moieties-glucose moiety], another key fragment ions of quercetin a glycone were observed at m/z 271 and 255. Therefore, compound 3 was identified as quercetin 3-(2 G-rhamnosylrutinoside) in comparison with the previously published data.[37] Compound 5 was detected at Rt = 16.18 min, it displayed a deprotonated molecule [M-H] at m/z 609, the neutral loss of rhamnosyl moiety (-m/z 146 u) afford a fragment ion at m/z 463 [M-H-rhamnose moiety], further loss of glucosyl moiety (-m/z 162 u) was confirmed through a diagnostic fragment ion at m/z 301 [M-H-rhamnosyl-glucosyl], which was assigned to quercetin a glycone and other key fragment ions of quercetin a glycone were detected at m/z 271, 255 and 179. Therefore, compound 5 was identified as quercetin-3-O-α-L-rhamnopyranosyl-(1 → 6)-β-D-glucopyranose (rutin).[36]

Compound 6 was detected at Rt = 17.18 min; it displayed a deprotonated molecule as a base peak at m/z 301, which was assigned to quercetin a glycone and other characteristic key fragment ions were appeared at m/z 271, 269, 255, 229, 179, and 151. Therefore, compound 6 was identified as 5, 7, 3′,4′-flavon-3-ol (quercetin).[38] Compound 7 was detected at Rt = 18.12 min, it displayed a deprotonated molecule [M-H] at m/z 463, the neutral loss of glucosyl moiety (-m/z 162 u) was confirmed through a fragment ion as a base peak at m/z 301 [M-H-glucose] was assigned to quercetin a glycone. Other key fragment ions were also observed at m/z 271, 255, 179, and 151. This fragmentation pattern was typically assigned to quercetin-3-O-β-D-glucoside (isoquercetin).[39],[40],[41] Compound 8 was detected at Rt = 19.29 min, it displayed a deprotonated molecule [M-H] at m/z 577, the neutral loss of rhamnosyl moiety (-m/z 146 u) give a fragment ion as a base peak at m/z 431 [M-H-rhamnosyl moiety] was assigned to kaempferol-O-rhamnoside, further neutral loss of the second rhamnosyl moiety (-m/z 146 u) was confirmed through the appearance of a fragment ion at m/z 285 which was attributed to kaempferol a glycone. Accordingly, compound 8 was identified as kaempferol-3,7-O-dirhamnoside (kaempferitrin).[40] Compound 9 was detected at Rt = 19.66 min, it displayed a deprotonated molecule [M-H] at m/z 623, the neutral loss of methyl moiety (-m/z 15 u) give a fragment ion at m/z 608 [M-H-CH3], while the neutral loss of rhamnosyl moiety (-m/z 146 u) give a fragment ion at m/z 477 [M-H-Rha], the loss of glucose moiety (-m/z 162 u) was confirmed through a diagnostic fragment ion as a base peak at m/z 315 [M-H-rutinoside], which was assigned to isorhamnetin a glycone. Therefore, compound 9 was identified as isorhamnetin-3-O-rutinoside.[40]

Compound 10 was detected at Rt = 20.18 min, it displayed a deprotonated molecule [M-H] at m/z 593, the neutral loss of rhamnosyl moiety (-m/z 146 u) give a fragment ion [M-H-Rha] at m/z 447 was assigned to kaempferol glucoside, further loss of the glucosyl moiety (-m/z 162 u) was confirmed through the appearance of a fragment ion as a base peak at m/z 285 [M-H-rhamnosyl-glucosyl] was assigned to kaempferol a glycone and due to total loss of rutinoside moiety. Therefore, compound 10 was identified as kaempferol-3-O-rutinoside.[41] Compound 11 was detected at Rt = 20.75 min, it displayed a deprotonated molecule [M-H] at m/z 447 was assigned to kaempferol glucoside, the neutral loss of glucosyl moiety (-m/z 162 u) give a fragment ion at m/z 285 [M-H-glucosyl] was assigned to kaempferol a glycone, and other key fragment ions were detected at m/z 255, 227, 179 and 151. Compound 11 could be identified as kaempferol-3-O-glucoside.[32],[39]

Compound 12 was detected at Rt = 21.19 min, it displayed a deprotonated molecule [M-H] at m/z 563, the neutral loss of rhamnosyl moiety (-m/z 146 u) give a fragment ion at m/z 417 [M-H-rhamnosyl], further loss of xylosyl moiety (-m/z 132) [M-H- rhamnosyl-xylosyl] was confirmed through the appearance of a diagnostic fragment ion at m/z 285 was assigned to kaempferol a glycone. Thus, compound 12 was tentatively identified as kaempferol-3-O-β-xylopyranosyl-(1→3)-α-L-rhamnopyranosyl. Compound 14 was detected at Rt = 22.67 min, it displayed a deprotonated molecule [M-H] at m/z 461, the loss of methyl moiety (-m/z 15 u) give a fragment ion at m/z 446 [M-H-CH3], while neutral loss of glucosyl moiety (-m/z 162 u) give a fragment ion as a base peak at m/z 299 [M-H-glu] was assigned to tri-hydroxy flavone nucleus. This fragmentation pattern was typically assigned to isoorientin-3′-O-methyl ether (isoscoparin).[42] Compound 20 was detected at Rt = 42.05 min, it displayed a deprotonated molecule [M-H] at m/z 329, the loss of methyl moiety (-m/z 15 u) give a fragment ion as a base peak at m/z 314 [M-H-CH3], further neutral loss of methyl moiety (-m/z 15 u) give a fragment ion at m/z 299 [M-H-2CH3], while the neutral loss of CO2 moiety (-m/z 44 u) give a fragment ion at m/z 285 [M-H-CO2], and another ion (-m/z 42 u) at m/z 243 [M-H-C2H2O-CO2], thus compound 20 could be identified as quercetin-dimethyl-ether.[43]

Compound 22 was detected at Rt = 49.38 min, the MS/Ms spectrum displayed a deprotonated molecular ion [M-H] at m/z 313, while the loss of methyl moiety (-m/z 15 u) give a fragment ion as a base peak at m/z 298 [M-H-CH3], further neutral loss of methyl moiety (-m/z 15 u) give a fragment ion at m/z 283 [M-H-2CH3], therefore, compound 22 was identified as 3′,7-dimethoxyluteolin (4′,5-dihydroxy-3′,7-dimethoxyflavone).[44] Compound 23 was detected at Rt = 51.05 min, the MS/Ms spectrum displayed a deprotonated molecular ion [M-H] at m/z 343, the loss of methyl moiety (-m/z 15 u) give a fragment ion as a base peak at m/z 328 [M-H-CH3], further neutral loss of methyl moiety (-m/z 15 u) give a fragment ion at m/z 313 [M-H-2CH3], other fragment ions were observed at m/z 297 and 285. Therefore, compound 23 was identified as dihydroxy-trimethoxy flavonol.[45]

Chalcones

Compound 13 was detected at Rt = 21.98 min, it displayed a deprotonated molecule [M-H] at m/z 391. The neutral loss of methyl moiety (-m/z 15 u) give a fragment ion as a base peak at m/z 376 [M-H-CH3] was assigned to methylated chalcone, the appearance of a fragment ion at m/z 135 was assigned to p-hydroxycinnamic acid moiety and due to the loss of 2,6-octa-dienyll-4-hydroxyphenyl moiety. Therefore, compound 13 was tentatively identified as 3'-(3,7-dimethyl-2,6-Octadienyl)-2'-4,4'-trihydroxy chalcone.[46]

Other detected compounds

Compound 18 was detected at Rt = 34.23 min, it displayed a deprotonated molecular ion [M-H] at m/z 327, the neutral loss of water moiety (-m/z 18 u) give a fragment ion at m/z 309 [M-H-H2O], the appearance of another fragment ion at m/z 291 due to further loss of another water moiety [M-H-2H2O], other fragment ions were observed at m/z 239, 229, 221, 211 and 171. Therefore, compound 18 was tentatively identified as 1,7-bis-(3,4-dihydroxyphenyl)-4-hepten-3-one (hirsutenone) in comparison with the previously published data.[47]

Phenolic compounds were detected in the n-BuOH extract

Phenolic acids and their derivatives

Compound 1 was detected at Rt = 2.76 min, it showed a deprotonated molecular ion [M-H] at m/z 315, a diagnostic fragment ion as a base peak was detected at m/z 153 due to the loss of glucosyl moiety (-m/z 162 u) and corresponding to protocatechuic acid [M-H-glucosyl moiety], further loss of CO2 moiety (-m/z 44 u) was confirmed through the appearance of fragment ion at m/z 109 [M-H-glucosyl moiety-CO2]. Therefore, compound 1 could be identified as protocatechuic acid glucoside.[36] Compound 14 was detected at Rt = 42.08 min, it showed a deprotonated molecular ion [M-H] at m/z 311, it also generated a fragment ion at m/z 179 corresponding to the loss of tartaric moiety (-m/z 132) [M-H-tartaric moiety], another fragment ion was appeared at m/z 149 due to the loss of caffeoyl moiety (-m/z 162) [M-H-caffeoyl moiety], this fragmentation profile was assigned to monocaffeyltartaric acid (caftaric).[35],[36]

Flavonoids

Compound 3 was detected at Rt = 14.70 min, it showed a deprotonated molecular ion [M-H] at m/z 623, it also generated a fragment ion at m/z 608 corresponding to the loss of methyl moiety (-m/z 15 u) [M-H-CH3], another fragment ion was observed at m/z 477 refer to the loss of rhamnosyl moiety (-m/z 146 u) [M-H-rhamnosyl moiety], it also generated a diagnostic fragment ion as a base peak at m/z 315 refer to further loss of glucosyl moiety (-m/z 162 u), which may represent isorhamnetin a glycone [M-H-rhamnosyl moiety-glucosyl moiety], another fragment ion was detected at m/z 300 due to the loss of methyl moiety from the a glycone nucleus [M-H-rhamnosyl moiety-glucosyl moiety-CH3], this fragmentation was typically assigned to isorhamnetin 3-O-rutinoside.[40] Compound 4 was detected at Rt = 15.04 min, it showed a deprotonated molecule [M-H] at m/z 755, a MS/MS fragment was appeared at m/z 609 corresponding to rutin molecule after the neutral loss of one rhamnosyl moiety (-m/z 146 u) [M-H-rhamnose moiety], a fragment ion was observed at m/z 445 due to the loss of another rhamnosyl and water moieties (-m/z 146-18 u) [M-H-2 rhamnose moiety-H2O], a diagnostic fragment ion was detected at m/z 301 was assigned to quercetin a glycone and can be explained by the release of the glucose moiety (-m/z 162 u) [M-H-2 rhamnose moieties-glucose moiety], in addition to the appearance of key fragment ions of quercetin a glycone at m/z 271, and 255. Therefore, compound 4 was identified as quercetin 3-(2 G-rhamnosylrutinoside).[37]

Compound 5 was detected at Rt = 15.80 min, it showed a deprotonated molecule [M-H] at m/z 739, a MS/MS fragment ion was observed at m/z 593 due to the loss of rhamnosyl moiety (-m/z 146 u) [M-H-rhamnose moiety], further neutral loss of water molecule (-m/z 18 u) led to generation of a fragment ion at m/z 575 [M-H-rhamnose moiety-H2O], further neutral loss of another rhamnosyl moiety (-m/z 146 u) led to generation of a fragment ion at m/z 429 [M-H-2rhamnose moiety-H2O], the appearance of fragment ion at m/z 285 was accounted for neutral loss of glucosyl moiety (-m/z 162 u) [M-H-2 rhamnose moiety-glucosyl moiety] it was assigned to kaempferol a glycone with key fragment ions at m/z 257, 255, and 227. Therefore, compound 5 was identified as kaempferol 3-O-rutinoside-7-O-rhamnoside.[41] Compound 8 was detected at Rt = 18.18 min, it showed a deprotonated molecule [M-H] at m/z 463, a diagnostic MS/MS fragment ion as a base peak was detected at m/z 301 due to the loss of glucosyl moiety (-m/z 162 u) [M-H-glucosyl moiety] it was assigned to quercetin a glycone, another key a glycone fragments were appeared at m/z 271, 255, 179 and 151. Thus compound 8 could be characterized as quercetin-3-O-β-D-glucoside.[39],[40],[41]

Compound 9 was detected at Rt = 19.95 min, it showed a deprotonated molecule [M-H] at m/z 593, a fragment ion was observed at m/z 447 refer to the release of rhamnosyl moiety (-m/z 146 u) [M-H-rhamnosyl moiety], further elimination of water molecule (-m/z 18 u) led to generation of fragment ion at m/z 429 [M-H-rhamnosyl moiety-H2O], while the release of glucosyl moiety (-m/z 162 u) led to generation of a diagnostic MS/MS fragment ion as a base peak at m/z 285 [M-H-rhamnosyl moiety-glucosyl moiety] it was assigned to kaempferol a glycone, another key a glycone fragments were detected at m/z 255, 227, 179, and 169. Therefore, compound 9 was identified as kaempferol 3-O- rutinoside.[41]

Compound 11 was detected at Rt = 21.80 min, it showed a deprotonated molecule [M-H] at m/z 447, while the release of glucosyl moiety (-m/z 162 u) led to generation of a diagnostic MS/MS fragment ion as a base peak at m/z 285 [M-H-glucosyl moiety] it was assigned to kaempferol a glycone, another key a glycone fragments were detected at m/z 255, 227, 179, 169 and 151. Therefore, compound 11 was identified as kaempferol-3-O-β-D-glucoside.[39],[42],[48] Compound 12 was detected at Rt 27.82 min, it showed a deprotonated molecule [M-H] at m/z 609, while the release of rhamnosyl moiety (-m/z 146 u) led to generation of MS/MS fragment ion at m/z 463 [M-H-rhamnosyl moiety] it was assigned to quercetin-O-glucoside, the removal of water molecule (-m/z 18 u) produce a fragment ion at m/z 445, while the elimination of the glucosyl moiety (-m/z 162 u) was confirmed through a diagnostic fragment at m/z 301 was assigned to quercetin a glycone, another key a glycone fragments were observed at m/z 271, 255, and 179. Therefore, compound 12 was identified as quercetin-3-O-rutinoside (rutin).[36] Compound 15 was detected at Rt = 42.90 min, it showed a deprotonated molecule [M-H] at m/z 329, while the release of methyl moiety (-m/z 15 u) led to generation of MS/MS fragment ion at m/z 314 [M-H-CH3] it was assigned to quercetin methyl ether, the removal of another methyl moiety (-m/z 15 u) produce a fragment ion at m/z 299 [M-H-2CH3]. Therefore, compound 15 was identified as quercetin-dimethyl-ether as previously reported.[43]

Phenylethanoid derivatives

Compound 13 was detected at Rt = 32.20 min, it showed a deprotonated molecule [M-H] at m/z 685, while the release of glucosyl moiety (-m/z 162 u) led to generation of a diagnostic MS/MS fragment ion as a base beak at m/z 523 [M-H-glucosyl moiety] it was assigned to ligstroside molecule, further removal of water molecule (-m/z 18 u) produce a fragment ion at m/z 505 [M-H-glucosyl moiety-H2O] and the elimination of another glucosyl moiety (-m/z 162 u) was confirmed through the appearance of a diagnostic fragment at m/z 343 [M-H-H2O-2glucosyl moiety]. Therefore, compound 13 was identified as ligstroside glucoside as previously described.[34]


   Conclusion Top


In the current study, the HPLC-DAD-ESI-MS/MS analysis led to the tentative identification of a total 30 phenolic compounds in the ethyl acetate and n-butanol extracts of S. nicolai leaves based on the determination of the precise mass of the deprotonated ions [M-H], which was obtained from the MS data and MSn fragmentation pattern. Among the tentatively identified compounds, flavonoids were the major constituents. To the best of our knowledge, this research work is the first comprehensive study on the polyphenolic composition of the Egyptian S. nicolai species. Moreover, S. nicolai extracts demonstrated considerable antioxidant and anticancer activities. S. nicolai could be further studied to isolate its biologically-active constituents and to study their modes of actions.

Acknowledgment

The authors wish to express their gratitude to Dr. Tearse Labib, Botany Specialist, Department of Flora and Taxonomy, El-Orman Botanical Garden, Giza, Egypt, for identification and authentication of the plant. Also, we would like to thank Dr. Mansour Sobeh & Dr. Tamer Mohamed; Department of Pharmaceutical Biology, Institute of Pharmacy and Molecular Biotechnology, Heidelberg University, Heidelberg, Germany, for their kind cooperation to perform LC/MS analysis.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
   References Top

1.
Hölscher D, Schneider B. Phenalenones from Strelitzia reginae. J Nat Prod 2000;63:1027-8.  Back to cited text no. 1
    
2.
Dwarka D, Thaver V, Naidu M, Baijnath H. New insights into the presence of bilirubin in a plant species Strelitzia nicolai (strelitziaceae). Afr J Tradit Complement Altern Med 2017;14:253-62.  Back to cited text no. 2
    
3.
Chalannavar RK, Venugopala KN, Baijnath H, Odhav B. Chemical composition of essential oil from the seed arils of Strelitzia nicolai regel and Koern from South Africa. J Essent Oil Bearing Plants 2014;17:1373-7.  Back to cited text no. 3
    
4.
Pirone C, Quirke JM, Priestap HA, Lee DW. Animal pigment bilirubin discovered in plants. J Am Chem Soc 2009;131:2830.  Back to cited text no. 4
    
5.
Pirone C, Johnson JV, Quirke JM, Priestap HA, Lee D. The animal pigment bilirubin identified in Strelitzia reginae, the bird of paradise flower. Hort Sci. 2010;45:1411-5.  Back to cited text no. 5
    
6.
Pirone C. Bilirubin: An Animal Pigment in the Zingiberales and Diverse Angiosperm Orders. Dissertation, Florida International University, UAS; 2010.  Back to cited text no. 6
    
7.
Dwarka D, Thaver V, Naidu M, Koorbanally NA, Baijnath AH. In vitro chemo-preventative activity of Strelitzia nicolai aril extract containing bilirubin. Afr J Tradit Complement Altern Med 2017;14:147-56.  Back to cited text no. 7
    
8.
Ghareeb MA, Saad AM, Abdou AM, Refahy LA, Ahmed WS. A new kaempferol glycoside with antioxidant activity from Chenopodium ambrosioides growing in Egypt. Orient J Chem 2016;32:3053-61.  Back to cited text no. 8
    
9.
Mangal M, Sagar P, Singh H, Raghava GP, Agarwal SM. NPACT: Naturally occurring plant-based anti-cancer compound-activity-target database. Nucleic Acids Res 2013;41:D1124-9.  Back to cited text no. 9
    
10.
Cheung-Ong K, Giaever G, Nislow C. DNA-damaging agents in cancer chemotherapy: Serendipity and chemical biology. Chem Biol 2013;20:648-59.  Back to cited text no. 10
    
11.
Ibrahim RM, El-Halawany AM, Saleh DO, El Naggar EB, El-Shabrawy AO, El-Hawary SS. HPLC-DAD-MS/MS profiling of phenolics from Securigera securidaca flowers and its anti-hyperglycemic and anti-hyperlipidemic activities. Rev Bras Farmacogn 2015;25:134-41.  Back to cited text no. 11
    
12.
Ghareeb MA, Mohamed T, Saad AM, Refahy LA, Sobeh M, Wink M. HPLC-DAD-ESI-MS/MS analysis of fruits from Firmiana simplex (L.) and evaluation of their antioxidant and antigenotoxic properties. J Pharm Pharm 2018;70:133-142.  Back to cited text no. 12
    
13.
Yang H, Dong Y, Du H, Shi H, Peng Y, Li X, et al. Antioxidant compounds from propolis collected in Anhui, China. Molecules 2011;16:3444-55.  Back to cited text no. 13
    
14.
Qader SW, Abdulla MA, Chua LS, Najim N, Zain MM, Hamdan S, et al. Antioxidant, total phenolic content and cytotoxicity evaluation of selected Malaysian plants. Molecules 2011;16:3433-43.  Back to cited text no. 14
    
15.
Goodman J, Chandna A, Roe K. Trends in animal use at US research facilities. J Med Ethics 2015;41:567-9.  Back to cited text no. 15
    
16.
Soliman RH, Ismail OA, Badr MS, Nasr SM. Resveratrol ameliorates oxidative stress and organ dysfunction in Schistosoma mansoni infected mice. Exp Parasitol 2017;174:52-8.  Back to cited text no. 16
    
17.
Dunham JH, Guthmiller P. Doing good science: Authenticating cell line identity. Cell Notes 2008;22:15-7.  Back to cited text no. 17
    
18.
Li L, Sharma N, Chippada U, Jiang X, Schloss R, Yarmush ML, et al. Functional modulation of ES-derived hepatocyte lineage cells via substrate compliance alteration. Ann Biomed Eng 2008;36:865-76.  Back to cited text no. 18
    
19.
Nasr SM, Ghareeb MA, Mohamed MA, Elwan NM, Abdel-Aziz AA, Abdel-Aziz MS. HPLC-Fingerprint analyses, in vitro cytotoxicity, antimicrobial and antioxidant activities of the extracts of two Cestrum species growing in Egypt. Pharmacog Res 2018;10:173-180.  Back to cited text no. 19
    
20.
S'liwka L, Wiktorska K, Suchocki P, Milczarek M, Mielczarek S, Lubelska K, et al. The comparison of MTT and CVS assays for the assessment of anticancer agent interactions. PLoS One 2016;11:e0155772.  Back to cited text no. 20
    
21.
Pietta PG. Flavonoids as antioxidants. J Nat Prod 2000;63:1035-42.  Back to cited text no. 21
    
22.
Arora A, Nair MG, Strasburg GM. Structure-activity relationships for antioxidant activities of a series of flavonoids in a liposomal system. Free Radic Biol Med 1998;24:1355-63.  Back to cited text no. 22
    
23.
Rice-Evans CA, Miller NJ, Paganga G. Structure-antioxidant activity relationships of flavonoids and phenolic acids. Free Radic Biol Med 1996;20:933-56.  Back to cited text no. 23
    
24.
Rice-Evans CA, Miller N, Paganga G. Antioxidant properties of phenolic compounds. Trends Plant Sci 1997;2:152-9.  Back to cited text no. 24
    
25.
Discher DE, Janmey P, Wang YL. Tissue cells feel and respond to the stiffness of their substrate. Science 2005;310:1139-43.  Back to cited text no. 25
    
26.
Sobeh M, Mahmoud MF, Hasan RA, Abdelfattah MAO, Sabry OM, Ghareeb MA, et al. Tannin-rich extracts from Lannea stuhlmannii and Lannea humilis (Anacardiaceae) exhibit hepatoprotective activities in vivo via enhancement of the anti-apoptotic protein Bcl-2. Sci Rep 2018;8:9343.  Back to cited text no. 26
    
27.
Rashdan HR, Nasr SM, El-Refai HA, Abdel-Aziz MS. A novel approach of potent antioxidant and antimicrobial agents containing coumarin moiety accompanied with cytotoxicity studies on the newly synthesized derivatives. J Appl Pharm Sci 2017;7:186-96.  Back to cited text no. 27
    
28.
Tanaka T, Kojima T, Kawamori T, Wang A, Suzui M, Okamoto K, et al. Inhibition of 4-nitroquinoline-1-oxide-induced rat tongue carcinogenesis by the naturally occurring plant phenolics caffeic, ellagic, chlorogenic and ferulic acids. Carcinogenesis 1993;14:1321-5.  Back to cited text no. 28
    
29.
Lee IR, Yang MY. Phenolic compounds from Duchesnea chrysantha and their cytotoxic activities in human cancer cell. Arch Pharm Res 1994;17:476-9.  Back to cited text no. 29
    
30.
Hertog MG, Kromhout D, Aravanis C, Blackburn H, Buzina R, Fidanza F, et al. Flavonoid intake and long-term risk of coronary heart disease and cancer in the seven countries study. Arch Intern Med 1995;155:381-6.  Back to cited text no. 30
    
31.
Yáñez J, Vicente V, Alcaraz M, Castillo J, Benavente-García O, Canteras M, et al. Cytotoxicity and antiproliferative activities of several phenolic compounds against three melanocytes cell lines: Relationship between structure and activity. Nutr Cancer 2004;49:191-9.  Back to cited text no. 31
    
32.
Abliz G, Mijit F, Hua L, Abdixkur G, Ablimit T, Amat N, et al. Anti-carcinogenic effects of the phenolic-rich extract from abnormal Savda Munziq in association with its cytotoxicity, apoptosis-inducing properties and telomerase activity in human cervical cancer cells (SiHa). BMC Complement Altern Med 2015;15:23.  Back to cited text no. 32
    
33.
Abu-Reidah IM, Arráez-Román D, Segura-Carretero A, Fernández-Gutiérrez A. Extensive characterisation of bioactive phenolic constituents from globe artichoke (Cynara scolymus L.) by HPLC-DAD-ESI-QTOF-MS. Food Chem 2013;141:2269-77.  Back to cited text no. 33
    
34.
Sanz M, de Simón BF, Cadahía E, Esteruelas E, Muñoz AM, Hernández T, et al. LC-DAD/ESI-MS/MS study of phenolic compounds in Ash (Fraxinus excelsior L. and F. americana L.) heartwood. Effect of toasting intensity at cooperage. J Mass Spectrom 2012;47:905-18.  Back to cited text no. 34
    
35.
Chen HJ, Inbaraj BS, Chen BH. Determination of phenolic acids and flavonoids in Taraxacum formosanum kitam by liquid chromatography-tandem mass spectrometry coupled with a post-column derivatization technique. Int J Mol Sci 2012;13:260-85.  Back to cited text no. 35
    
36.
Abu-Reidah IM, Ali-Shtayeh MS, Jamous RM, Arráez-Román D, Segura-Carretero A. HPLC-DAD-ESI-MS/MS screening of bioactive components from Rhus coriaria L. (Sumac) fruits. Food Chem 2015;166:179-91.  Back to cited text no. 36
    
37.
Abu-Reidah IM, Arráez-Román D, Lozano-Sánchez J, Segura-Carretero A, Fernández-Gutiérrez A. Phytochemical characterisation of green beans (Phaseolus vulgaris L.) by using high-performance liquid chromatography coupled with time-of-flight mass spectrometry. Phytochem Anal 2013;24:105-16.  Back to cited text no. 37
    
38.
Abu-Reidah IM, del Mar Contreras M, Arráez-Román D, Fernández-Gutiérrez A, Segura-Carretero A. UHPLC-ESI-QTOF-MS-based metabolic profiling of Vicia faba L. (Fabaceae) seeds as a key strategy for characterization in foodomics. Electrophoresis 2014;35:1571-81.  Back to cited text no. 38
    
39.
Bravo L, Goya L, Lecumberri E. LC/MS characterization of phenolic constituents of mate (Ilex paraguariensis, St. Hil.) and its antioxidant activity compared to commonly consumed beverages. Food Res Int 2007;40:393-405.  Back to cited text no. 39
    
40.
Abu-Reidah IM, Arráez-Román D, Quirantes-Piné R, Fernández-Arroyo S, Segura-Carretero A, Fernández-Gutiérrez A. HPLC–ESI-Q-TOF-MS for a comprehensive characterization of bioactive phenolic compounds in cucumber whole fruit extract. Food Res Int 2012;46:108-17.  Back to cited text no. 40
    
41.
Al-Rawahi AS, Edwards G, Al-Sibani M, Al-Thani G, Al-Harrasi AS, Rahman MS. Phenolic constituents of pomegranate peels (Punica granatum L.) cultivated in oman. Eur J Med Plants 2014;4:315-31.  Back to cited text no. 41
    
42.
Chen F, Long X, Liu Z, Shao H, Liu L. Analysis of phenolic acids of Jerusalem artichoke (Helianthus tuberosus L.) responding to salt-stress by liquid chromatography/tandem mass spectrometry. ScientificWorldJournal 2014;2014:568043.  Back to cited text no. 42
    
43.
Pellati F, Orlandini G, Pinetti D, Benvenuti S. HPLC-DAD and HPLC-ESI-MS/MS methods for metabolite profiling of Propolis extracts. J Pharm Biomed Anal 2011;55:934-48.  Back to cited text no. 43
    
44.
Simirgiotis MJ, Benites J, Areche C, Sepúlveda B. Antioxidant capacities and analysis of phenolic compounds in three endemic Nolana species by HPLC-PDA-ESI-MS. Molecules 2015;20:11490-507.  Back to cited text no. 44
    
45.
Abdel-Hameed ES, Bazaid SA, Shohayeb MM. RP-HPLC-UV-ESI-MS phytochemical analysis of fruits of Conocarpus erectus L. Chem Pap 2014;68:1358-67.  Back to cited text no. 45
    
46.
Kim DW, Curtis-Long MJ, Yuk HJ, Wang Y, Song YH, Jeong SH, et al. Quantitative analysis of phenolic metabolites from different parts of Angelica keiskei by HPLC-ESI MS/MS and their xanthine oxidase inhibition. Food Chem 2014;153:20-7.  Back to cited text no. 46
    
47.
Riethmüller E, Alberti A, Tóth G, Béni S, Ortolano F, Kéry A, et al. Characterisation of diarylheptanoid- and flavonoid-type phenolics in Corylus avellana L. Leaves and bark by HPLC/DAD-ESI/MS. Phytochem Anal 2013;24:493-503.  Back to cited text no. 47
    
48.
Ágnes A. LC-ESI-MS/MS methods in profiling of flavonoid glycosides and phenolic acids in traditional medicinal plants: Sempervivum tectorum L. and Corylus avellana L. Dissertation, Semmelweis University, Hungary; 2013.  Back to cited text no. 48
    


    Figures

  [Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6], [Figure 7]
 
 
    Tables

  [Table 1], [Table 2], [Table 3], [Table 4], [Table 5]



 

Top
  
 
  Search
 
    Similar in PUBMED
   Search Pubmed for
   Search in Google Scholar for
 Related articles
    Access Statistics
    Email Alert *
    Add to My List *
* Registration required (free)  

 
  In this article
    Abstract
   Introduction
    Materials and Me...
    Results and Disc...
   Conclusion
    References
    Article Figures
    Article Tables

 Article Access Statistics
    Viewed185    
    Printed14    
    Emailed0    
    PDF Downloaded5    
    Comments [Add]    

Recommend this journal