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Comparison of the polyphenolic profile and antibacterial activity of the leaves, fruits and flowers of Rhododendron ambiguum and Rhododendron cinnabarinum

BMC Research Notes volume 10, Article number: 297 (2017) Cite this article

Abstract

Background

Rhododendron species have been traditionally used in countries like China, Nepal, Russia and North America for treating human diseases. These species are known to be a good source of polyphenolic plant secondary plant metabolites. They are known to have beneficial health properties for humans and have been used to treat diseases like asthma, skin diseases. In this contribution we investigate the phenolic profile and antibacterial activity of extracts from several plant organs including for the first time from leaves of different development stages.

Methods

In this study, the polyphenolic profile of fruits, flowers and leaves of different ages of Rhododendron ambiguum and Rhododendron cinnabarinum were studied by using HPLC–MS and compounds identified based on high resolution masses and identity of tandem mass spectra, UV/VIS spectra and retention times if compared to standards.

Results

Fifty-nine different polyphenols including isomers were identified in these species by their fragmentation pattern and high resolution data. Also, the antibacterial activity of these parts (leaves, fruits and flowers) against gram-positive bacteria was studied.

Conclusion

The leaves and fruits contained more polyphenols than the flowers. With the exception of flowers, the fruits and leaves of both species were also determined to have a significant antibacterial effect against four gram-positive bacteria.

Background

Polyphenols are secondary plant metabolites that are known to have different physical, chemical and biological properties. The four major types of polyphenols are flavonoids, phenolic acids, lignans and stilbenes. The phenolic acid polyphenols consist of the derivatives of hydroxybenzoic acid and hydroxycinnamic acid. The flavonoids are further classified into different groups based on their structure variation and hydroxylation pattern. Proanthocyanidins (PAs) belong to the class of flavonoids, which are formed from the oligomerization and polymerization of flavan-3-ol units such as catechin and afzelechin (Fig. 1). PAs are commonly found in fruits, vegetables and grain [1, 2]. They are known to have anti-inflammatory [3] and antimutagenic [4] effects and have been used in the treatment of asthma [4], skin diseases and UV radiations [5].

Fig. 1
figure 1

Representative examples of polyphenols from Rhododendron species. For the complete list of structures of compounds identified, please refer to Additional file 1

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Rhododendron is a genus of woody plants that belong to Ericaceae family. Most of the species have attractive flowers and more than 1000 species of the genus have been described. They differ in their range of size, shape, texture, growth habit and color of blossoms [6]. The genus is ranging from shrubs and small to large trees. Rhododendron species have decorative flowers, which are mainly used for ornamental purposes. There is variation in the height of the plant, starting from 10 cm to 1 m (smallest species), while the example of largest specie is R. giganteum, which is almost 3 m tall. The leaves of most species are spirally arranged; the nature of leaves may be evergreen or deciduous.

The genus Rhododendron is found in almost all parts of the world except some parts in America and Africa. Species of the genus Rhododendron occur throughout the Northern Hemisphere and the Southern Hemisphere in South Eastern Asia and Northern Australasia. The plant is originally present in mountainous areas characterized by acidic well-drained soil, regular rainfall and cool summer temperatures [7]. On the other hand, majority of the genus prefers cooler temperature including the subgenus Hymenanthes. One of the subsections (Pontica) is indigenous to the areas outside the center of distribution and present in the areas of Japan, eastern China, Europe, North America and Russia [8].

Rhododendron (flowers and leaves) has traditionally been used in China, Nepal, North America, Russia, Korea, Austria and Romania for treating various diseases including arthritis, intestinal disorders, rheumatism, skin diseases, cough and other ailments [9]. They are known to be rich in polyphenolic compounds namely flavonoids and their glycosides, terpenoids and essential oils [1].

The new and exciting aspect of this study is the analysis of different plant organs where only limited data is available in the literature. The hypotheses of this study are that fruits, flowers and leaves have different polyphenolic profile and that the leaves of different ages, which are exposed to numerous environmental conditions might lead to the production of new compounds or variation in secondary metabolite quantities. In particular Rhododendron as an evergreen shrub grows a new generation of leaves every year offering the unique opportunity to compare the polyphenolic profile of leaves of different ages and hence study the biochemical history of the plant. Such an investigation has to the best of our knowledge never been carried out. The polyphenolic profile of the leaves of different age, flowers and fruits were analyzed for R. ambiguum and R. cinnabarinum. Both species belong to the subgenus Rhododendron, section Rhododendron. R. ambiguum and R. cinnabarinum belong to the subsection Triflora and Cinnabarina respectively. R. cinnabarinum is considered to be toxic for animals [9, 10]. It is known from our previous studies that these two species out of 17 Rhododendron species showed higher activity against several Gram-positive bacteria [11]. Moreover, the high dose of both Rhododendron species exhibited a toxic effect in two mammalian cells and induced phenotypic changes that are characteristic for apoptosis [12]. Thus, the aim of the study was to analyze the chemical profile of Rhododendron crude extracts in order to contribute to our on-going investigations on the bioactivity potential of Rhododendron.

Methods

Plant material and chemicals

Fresh leaf material of R. ambiguum Hemsley and R. cinnabarinum Hooker (first, second, and third leaf) were collected from plants grown in the Rhododendron-Park Bremen (http://www.rhododendronparkbremen.de) from April 2013 at 10:00 [gene bank number: R. ambiguum (100.007); R. cinnabarinum (100.322)]. First and second year leaves were collected for R. ambiguum, and first, second and third year leaves were collected for R. cinnabarinum. The leaves were distinguished on the basis of their morphological features. Moreover, the flowers and fruits for both Rhododendron species were also sampled. Each sample species was collected from three different individual plants with the help of Dr. Hartwig Schepker. The identities of all plant species have been authenticated according to the German Genebank Rhododendron Database provided by the Bundessortenamt (http://www.bundessortenamt.de/rhodo). Samples were deposited in herbaria with voucher numbers: R. ambiguum: OLD00801; R. cinnabarinum: OLD00757.

All chemicals (analytical grade) were purchased from Carl Roth (Karlsruhe, Germany). Standards were used whenever applicable, to compare the fragmentation and retention time of the compounds.

Plant extraction

The Rhododendron leaves, fruits and flowers were freeze dried using liquid nitrogen. A mortar and pestle was used to crush the dried brittle leaves. 2 g of the powdered material were dissolved in 10 mL of 80% aqueous methanol and for 24 h at 4 °C. The mixture was then sonicated for 15 min and centrifuged at 3.220×g for 10 min. The aliquot was then separated and stored at −20 °C until further analyses.

LC-ESI-TOF–MS (high resolution mass spectrometry)

The LC equipment (Agilent 1200 series, Bremen, Germany) consists of a binary pump, an auto-sampler with 100 µL loop and a UV–Vis detector with a light-pipe flow cell. The UV detector was used at 280 nm to measure the polyphenols. The 5 µm diphenyl column having 250 × 3 mm inner diameter (Varian, Darmstadt, Germany) was used for separation. This was connected to the microTOF mass spectrometer (Bruker Daltonics, Bremen, Germany) equipped with an electrospray ionization source. The internal calibration was achieved by using 0.1 M sodium formate solution at 0.10 mL/min, which was injected through the six-port valve. The calibration was achieved by using the enhanced quadratic mode. Water/formic acid (1000:0.05 v/v) and methanol were used as solvent A and B respectively. The flow rate of the solvents was adjusted to 500 µL/min. A linear gradient was used from 10% B to 80% B in 70 min and a further 10 min was assigned for the gradient to equilibrate from 80% B to 10% B for the next run. 3 µL of the filtered extract was injected into the system. The software used in this system was Bruker Hystar.

LC-ESI-MSn (Tandem mass spectrometry)

The Liquid chromatography equipment (Agilent 1100 series) comprises of a binary pump, an auto sampler having a 100 µL capacity loop and a Diode Array Detector with a range from 200 to 600 nm. The detector recorded at 254, 280 and 320 nm, which is the best absorption wavelength for polyphenolic compounds. Chromatographic separation was performed using the same gradient method used in the LC-TOF analyses. A 5 μm diphenyl column of 250 × 3 mm i.d. (Varian, Darmstadt, Germany) with 500 µL/min flow rate of solvent was used. The LC equipment was connected with Ion-trap mass spectrometer, which was fitted with an ESI source (Bruker Daltonics HCT Ultra, Bremen, Germany) operating in full scan auto MSn mode to obtain fragment ions. Tandem mass spectra were acquired in Auto-MSn mode (smart fragmentation) using a ramping of the collision energy. Maximum fragmentation amplitude was fixed to 1 V. MS operating conditions (negative mode) had been optimized with a capillary temperature of 365 °C, a dry gas flow rate was of 10 L/min, and a nebulizer pressure of 10 psi. 3 µL of the filtered extract was injected into the system. The software used in this system was Agilent Chemstation.

Bacterial strains and antimicrobial susceptibility test

Four Gram-Positive bacterial species i.e. Bacillus subtilis S168, Bacillus aquimaris MB-2011, Bacillus thioparus, and Clavibacter michiganensis and one Gram-Negative bacterial specie i.e. Escherichia coli were selected to compare the susceptibility of crude extracts of leaves, flower and fruit of two Rhododendron species i.e. R. ambiguum and R. cinnabarinum. Antimicrobial activity screening was conducted by the agar diffusion method [13]. Briefly, Lysogeny Broth (LB) agar plates were inoculated with 200 µL of the inoculum of the tester organism (1 × 107 colony forming units per mL) by evenly spreading the cell suspensions over the agar surface. Holes with diameters of 5 mm were punched into the agar plates. Subsequently, 50 µL of the plant crude extracts were filled into each well. The plates were incubated overnight at 28 °C. Inhibition of microbial growth was determined by measuring the radius of the inhibition zone. For each bacterial strain, 80% aqueous methanol solutions were used as negative solvent controls. All experiments were performed in triplicates and the results were presented as mean values. Ampicillin was used as the positive control and the extraction solvent, 80% aqueous methanol was used as the negative control.

Results and discussion

To our knowledge, no investigation on the polyphenolic profile of leaves of different ages has been carried out. The crude extracts of all parts of the two selected species R. ambiguum and R. cinnabarinum (i.e. first, second, third leaves, flowers and fruits) were extracted with 80% aqueous methanol in order to extract polyphenols. Both species were recently shown to possess promising biological activities [11] and their plants consisted of leaves of different age. These extracts were analyzed by reversed phase HPLC using a diphenyl column with a gradient using methanol and water/formic acid (1000:0.05 v/v). Negative ion mode was used to study the polyphenols using tandem mass spectrometry and high resolution mass spectrometry. The compounds were identified in the high resolution mass data by observing an absolute mass error below 5 ppm for their elemental composition. The UV spectrum at 280 nm was used to identify the PAs present in the samples. Also, the fragmentation pathway of PAs by heterocyclic ring fission (HRF) and retro-Diels–Alder (RDA) reaction were considered [14].

Total ion chromatograms from LC–MS

The chromatograms of Rhododendron cinnabarinum first year leaves, second year leaves, third year leaves, flowers and fruits were directly compared and are shown in Fig. 2. The chromatograms of the first, second and third year leaves consisted of peaks at identical retention time and comparable intensities. The chromatograms of flowers and fruits were different compared to the leaves. The chromatogram of flowers consisted of peaks with lower intensity in the region 0–30 min, which suggests that the hydrophilic compounds are present in lower intensity compared to the leaves. However, after 30 min, the chromatogram of the flowers was similar to the leaves. On the other hand, the chromatogram of the fruits exhibited additional peaks not present in the leaves and flowers.

Fig. 2
figure 2

Total Ion Chromatogram of Rhododendron cinnabarinum leaves, flowers and fruits by LC-MSn in negative mode

Full size image

In this study, 59 polyphenolic compounds were identified from both Rhododendron species for the first, second third year leaves, flower and fruit (Table 1). The identification of reported compounds was based on their retention time, fragmentation pattern and high resolution mass data. All identified polyphenol compounds have been discovered in nature already [1, 15,16,17,18,19,20,21,22,23,24,25,26]. Out of the identified compounds, 46 were already reported to be found in Rhododendron species [1, 16,17,18]. In this study, 13 new polyphenolic compounds were identified in Rhododendron species. The fruits consisted of the highest variety and concentration of PAs among the four parts of R. ambiguum and R. cinnabarinum.

Table 1 Polyphenols present in different year leaves, flowers and fruits of R. ambiguum and R. cinnabarinum
Full size table

Characterization of methyl gallate hexoside [1] (Mr 345)

A peak was detected at m/z 345 and was assigned to be methyl gallate hexoside. The peak produced the fragmentation with base peak at m/z 183 and secondary peaks of m/z 168 and 124.

Characterization of (epi)catechin-(4,8′/2,6′)-(epi)catechin [53, 54] (Mr 576)

Two peaks were detected at m/z 575 and were assigned as A-type dimer of (epi)catechin unit. The first peak produced an MS2 base peak of m/z 449 and secondary peak of m/z 287. The second peak produced an MS2 base peak of m/z 423 and secondary peak of m/z 285.

Characterization of catechin [55] and epicatechin [56] (Mr 290)

Two peaks were detected at m/z 289. The first peak was assigned as catechin and the second peak was assigned as epicatechin based on their polarity. Both peaks produced similar fragmentation, consisting of an MS2 base peak of m/z 245 and secondary peak of m/z 203.

Characterization of gallocatechin [57] and epigallocatechin [58] (Mr 306)

Two peaks were detected at m/z 305. The first peak was assigned as gallocatechin and the second peak was assigned as epigallocatechin based on their polarity. Both peaks produced similar fragmentation, consisting of an MS2 base peak of m/z 179 and secondary peak of m/z 164.

Characterization of (epi)catechin-(4,8′)-(epi)catechin [39, 40, 41, 42, 43] (Mr 578)

Five peaks were detected at m/z 577. They were assigned as the PA dimer. All the peaks produced similar fragmentation with the base peak of m/z 407 and secondary peaks of m/z 425 and 285. The standards of the dimers B1 and B2 were used to differentiate the isomers by their retention time.

Characterization of (epi)gallocatechin-(4,8′)-(epi)catechin and (epi)catechin-(4,8′)-(epi)gallocatechin [47, 48, 49, 50, 51, 52] (Mr 594)

Six peaks were detected at m/z 593. They were speculated to be dimeric B-type PA consisting of (epi)catechin and (epi)gallocatechin monomeric units. All the peaks produced similar fragmentation having the base peak of m/z 425 and secondary peak of m/z 407.

Characterization of (epi)gallocatechin-(4,8′)-(epi)gallocatechin [36, 37, 38] (Mr 610)

Three peaks were detected at m/z 609 and were speculated to be dimeric B-type PAs with (epi)gallocatechin monomeric units. The three peaks produced the fragmentation with base peak at m/z 423 and secondary peaks of m/z 441 and 283.

Characterization of Taxifolin [27] (Mr 304)

A peak was detected at m/z 303 and was assigned to be taxifolin. The peak produced the fragmentation with base peak at m/z 285 and secondary peaks of m/z 177 and 125.

Identification of other polyphenols

The polyphenols were identified by their specific fragmentation patterns, retention time and high resolution mass values. The other polyphenols that were identified in the leaf extracts are three vanillic acid-O-hexosides [19], two salicylic acid hexosides [20, 28], three caffeoylquinic acids [1], naringenin [29], myricetin [29], two myricetin-O-hexosides [1], myricetin-O-rhamnoside [1], three myricetin-O-pentosides [1], two quercetin-O-hexosides [1], three quercetin-O-pentosides [1], quercetin-O-rhamnoside [1], quercetin-O-rhamnoside-O-hexoside [1], quercetin-O-glucoronide [21, 30], quercetin [1], kaempferol [27, 29], kaempferol-O-rhamnoside [22], two kaempferol-O-pentosides [22], kaempferol-O-glucoronide [23, 30], three taxifolin-O-pentosides [1, 31], two procyanidin trimers C [25, 32], one A-type procyandintrimer C [21, 24] and (epi)catechin-O-d-glycopyranoside [25]. The tandem mass spectra of some compounds are shown in Fig. 3.

Fig. 3
figure 3

Tandem MS fragmentation of some polyphenols identified in Rhododendron

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Antibacterial activity

Four Gram-positive organisms were used to compare the antibacterial activity of different plant parts of two species from genus Rhododendron. Crude extract of first, second, third year leaves in addition to flowers and fruits of R. ambiguum and R. cinnabarinum were obtained using 80% methanol. There was no bioactivity observed for crude extracts of all samples against Escherichia coli. Against Gram-positive bacteria, the bioactivity of R. ambiguum ranged between 0.5 and 0.7 cm, while for R. cinnabarinum between 0.5 and 0.8 cm (Fig. 4). Antibacterial effects of fruit and leaves extracts were in the same order of magnitude. However, there was a reduced antibacterial activity observed for the flowers of R. cinnabarinum and R. ambiguum. This could be due to the evolutionary aspect as flowers have a short blooming period in a year compared to the leaves and fruits. In general, B. thioparus was the most sensitive bacteria species towards the plant parts for both Rhododendron species. These results are in agreement with our previous study, which showed a higher antibacterial effect of Rhododendron species against Gram-positive and higher effect for R. cinnabarinum [11]. The polyphenolic analysis indicated that Taxifolin derivatives were present in high concentration in the leaves of both plant species, which could be the reason of apoptosis like phenotype observed before in other studies [12]. Moreover, this finding is also supported by other studies, which reported the effect of Taxifolin in different cancer cell lines by inducing apoptosis cell death [33, 34].

Fig. 4
figure 4

Antimicrobial activities of methanol crude extract of different plant parts. a R. ambiguum and b R. cinnabarinum

Full size image

The radius of the inhibition zones was measured in triplicates and the values are given as means ± standard deviations. The aqueous methanol used as negative controls did not yield inhibition zones (data not shown).

Since flowers were found to have lower bioactivity as compared to the leaves and fruits, a list of compounds present in either leaves or fruits, but not in flowers was compiled. This list consisted of 1 (methyl gallate hexoside), 6 (salicylic acid-O-hexoside), 9 (4-O-caffeoylquinic acid), 14 (myricetin-O-rhamnoside), 15, 16 (myricetin-O-pentoside), 20 (quercetin-O-pentoside), 24 (quercetin-O-rhamnoside-O-hexoside), 31 (kaempferol-3-O-glucuronide), 33, 35 (taxifolin-O-pentoside), 36, 37, 38 (epi)gallocatechin-(epi)gallocatechin, 44, 45 (procyanidin trimer C), 47, 49, 50, 52 ((epi)gallocatechin-(epi)catechin) and 59 ((epi)catechin-O-D-glucopyranoside). The presence of these compounds in the plant organs could be either a contributing factor to the bioactivity itself or could function as a marker for plant organs with potential bioactivity.

Conclusions

The different parts of the R. ambiguum and R. cinnabarinum are a rich source of polyphenols. Fifty-nine different types of polyphenols including isomers were identified in these parts based on their fragmentation pattern and high resolution mass spectra in negative ion mode. The polyphenolic profile of different year leaves was found to be similar. Among all the parts, the fruits were found to contain the highest variety and concentration of polyphenols. PAs were mainly found in the fruits. However, there are many unidentified compounds present in leaves, flowers and fruits, which need to be analyzed in future. We can conclude that both Rhododendron species have antibacterial effect towards Gram-positive bacteria, while there was no significant difference between different seasonal leaves and fruits, but low effect for flowers.

Abbreviations

ESI:

electro spray ionization

HPLC:

high performance liquid chromatography

LB:

Lysogeny Broth

LC:

liquid chromatography

m/z :

mass to charge ratio

M:

molar

Mr :

molecular weight

MS:

mass spectrometry

MSn :

tandem mass spectrometry

PAs:

proanthocyanidins

psi:

pound-force per square inch

RT:

retention time

TOF:

time of flight

UV:

ultra violet

v/v:

volume/volume

Vis:

visible

References

  1. Jaiswal R, Jayasinghe L, Kuhnert N. Identification and characterization of proanthocyanidins of 16 members of the Rhododendron genus (Ericaceae) by tandem LC–MS. J Mass Spectrom. 2012;47(4):502–15.

    Article  CAS  PubMed  Google Scholar 

  2. Kuhnert N, Karakoese H, Jaiswal R. Analysis of chlorogenic acids and other hydroxycinnamates in food, plants, and pharmacokinetic studies. In: Anonymous, editor. Boca Raton: CRC Press; 2012. p. 461–510.

  3. Kruger MJ, Davies N, Myburgh KH, Lecour S. Proanthocyanidins, anthocyanins and cardiovascular diseases. Food Res Int. 2014;59:41–52.

    Article  CAS  Google Scholar 

  4. Zhou DY, Du Q, Li RR, Huang M, Zhang Q, Wei GZ. Grape seed proanthocyanidin extract attenuates airway inflammation and hyperresponsiveness in a murine model of asthma by downregulating inducible nitric oxide synthase. Planta Med. 2011;77(14):1575–81.

    Article  CAS  PubMed  Google Scholar 

  5. Deters A, Dauer A, Schnetz E, Fartasch M, Hensel A. High molecular compounds (polysaccharides and proanthocyanidins) from Hamamelis virginiana bark: influence on human skin keratinocyte proliferation and differentiation and influence on irritated skin. Phytochemistry. 2001;58(6):949–58.

    Article  CAS  PubMed  Google Scholar 

  6. Dirr MA. Manual of woody landscape plants: their identification, ornamental characteristics, culture, propagation and uses. 4th ed. Champaign; 2009.

  7. Horn CN. Distribution and ecological preference of Rhododendron eastmanii Kron & Creel (May-White Azalea) in South Carolina. Castanea. 2005;70(1):1–12.

    Article  Google Scholar 

  8. Chamberlain DF, Tian-Lu M. The distribution of Rhododendron subgenus Hymenanthes with special reference to Yunnan (W China). Notes RBG Edinburgh. 1985;43(1):7–13.

    Google Scholar 

  9. Popescu R, Kopp B. The genus Rhododendron: an ethnopharmacological and toxicological review. J Ethnopharmacol. 2013;147(1):42–62.

    Article  CAS  PubMed  Google Scholar 

  10. Anonymous Rhododendron cinnabarinum Hook. In: Anonymous, editor. New York: Springer; 2007. p. 1.

  11. Rezk A, Nolzen J, Schepker H, Albach DC, Brix K, Ullrich MS. Phylogenetic spectrum and analysis of antibacterial activities of leaf extracts from plants of the genus Rhododendron. BMC Complement Altern Med. 2015;15:67.

    Article  PubMed  PubMed Central  Google Scholar 

  12. Rezk A, Schepker H, Brix K, Ullrich MS: Assessment of cytotoxicity exerted by leaf extracts from plants of the genus Rhododendron towards epidermal keratinocytes and intestine epithelial cells. Submitted 2015.

  13. Nathan P, Law EJ, Murphy DF, MacMillan BG. A laboratory method for selection of topical antimicrobial agents to treat infected burn wounds. Burns. 1978;4(3):177–87.

    Article  Google Scholar 

  14. Gu L, Kelm MA, Hammerstone JF, Zhang Z, Beecher G, Holden J, Haytowitz D, Prior RL. Liquid chromatographic/electrospray ionization mass spectrometric studies of proanthocyanidins in foods. J Mass Spectrom. 2003;38(12):1272–80.

    Article  CAS  PubMed  Google Scholar 

  15. Cserhati T, Forgacs E, Morais MH, Mota T. Liquid chromatography of natural pigments. Biomed Chromatogr. 2000;14(5):281–6.

    Article  CAS  PubMed  Google Scholar 

  16. Swiderski A, Muras P, Koloczek H. Flavonoid composition in frost-resistant Rhododendron cultivars grown in Poland. Sci Hortic. 2004;100(14):139–51.

    Article  CAS  Google Scholar 

  17. Cao Y, Chu Q, Ye J. Chromatographic and electrophoretic methods for pharmaceutically active compounds in Rhododendron dauricum. J Chromatogr B. 2004;812(12):231–40.

    Article  CAS  Google Scholar 

  18. Louis A, Petereit F, Lechtenberg M, Deters A, Hensel A. Phytochemical characterization of Rhododendron ferrugineum and in vitro assessment of an aqueous extract on cell toxicity. Planta Med. 2010;76(14):1550–7.

    Article  CAS  PubMed  Google Scholar 

  19. Vallverdu-Queralt A, Jauregui O, Di Lecce G, Andres-Lacueva C, Lamuela-Raventos RM. Screening of the polyphenol content of tomato-based products through accurate-mass spectrometry (HPLC–ESI-QTOF). Food Chem. 2011;129(3):877–83.

    Article  CAS  PubMed  Google Scholar 

  20. Abu-Reidah IM, Arraez-Roman D, Segura-Carretero A, Fernandez-Gutierrez A. Profiling of phenolic and other polar constituents from hydro-methanolic extract of watermelon (Citrullus lanatus) by means of accurate-mass spectrometry (HPLC-ESI-QTOF-MS). Food Res Int. 2013;51(1):354–62.

    Article  CAS  Google Scholar 

  21. Ma Y, Kosinska-Cagnazzo A, Kerr WL, Amarowicz R, Swanson RB, Pegg RB. Separation and characterization of phenolic compounds from dry-blanched peanut skins by liquid chromatography-electrospray ionization mass spectrometry. J Chromatogr A. 2014;1356:64–81.

    Article  CAS  PubMed  Google Scholar 

  22. Lhuillier A, Fabre N, Moyano F, Martins N, Claparols C, Fouraste I, Moulis C. Comparison of flavonoid profiles of Agauria salicifolia (Ericaceae) by liquid chromatography-UV diode array detection-electrospray ionisation mass spectrometry. J Chromatogr A. 2007;1160(1–2):13–20.

    Article  CAS  PubMed  Google Scholar 

  23. De Rosso M, Tonidandel L, Larcher R, Nicolini G, Dalla Vedova A, De Marchi F, Gardiman M, Giust M, Flamini R. Identification of new flavonols in hybrid grapes by combined liquid chromatography–mass spectrometry approaches. Food Chem. 2014;163:244–51.

    Article  PubMed  Google Scholar 

  24. Li S, Xiao J, Chen L, Hu C, Chen P, Xie B, Sun Z. Identification of A-series oligomeric procyanidins from pericarp of Litchi chinensis by FT-ICR-MS and LC-MS. Food Chem. 2012;135(1):31–8.

    Article  CAS  Google Scholar 

  25. Zhao T, He J, Wang X, Ma B, Wang X, Zhang L, Li P, Liu N, Lu J, Zhang X. Rapid detection and characterization of major phenolic compounds in Radix Actinidia chinensis Planch by ultra-performance liquid chromatography tandem mass spectrometry. J Pharm Biomed Anal. 2014;98:311–20.

    Article  CAS  PubMed  Google Scholar 

  26. Jaiswal R, Karakose H, Ruhmann S, Goldner K, Neumuller M, Treutter D, Kuhnert N. Identification of phenolic compounds in plum fruits (Prunus salicina L. and Prunus domestica L.) by high-performance liquid chromatography/tandem mass spectrometry and characterization of varieties by quantitative phenolic fingerprints. J Agric Food Chem. 2013;61(49):12020–31.

    Article  CAS  PubMed  Google Scholar 

  27. Sapozhnikova Y. Development of liquid chromatography-tandem mass spectrometry method for analysis of polyphenolic compounds in liquid samples of grape juice, green tea and coffee. Food Chem. 2014;150:87–93.

    Article  CAS  PubMed  Google Scholar 

  28. Torras-Claveria L, Jáuregui O, Codina C, Tiburcio AF, Bastida J, Viladomat F. Analysis of phenolic compounds by high-performance liquid chromatography coupled to electrospray ionization tandem mass spectrometry in senescent and water-stressed tobacco. Plant Sci. 2012;182:71–8.

    Article  CAS  PubMed  Google Scholar 

  29. Biesaga M, Pyrzynska K. Liquid chromatography/tandem mass spectrometry studies of the phenolic compounds in honey. J Chromatogr A. 2009;1216(38):6620–6.

    Article  CAS  PubMed  Google Scholar 

  30. Zhou H, Liang J, Lv D, Hu Y, Zhu Y, Si J, Wu S. Characterization of phenolics of Sarcandra glabra by non-targeted high-performance liquid chromatography fingerprinting and following targeted electrospray ionisation tandem mass spectrometry/time-of-flight mass spectrometry analyses. Food Chem. 2013;138(4):2390–8.

    Article  CAS  PubMed  Google Scholar 

  31. Ruiz A, Bustamante L, Vergara C, von Baer D, Hermosín-Gutierrez I, Obando L, Mardones C. Hydroxycinnamic acids and flavonols in native edible berries of South Patagonia. Food Chem. 2015;167:84–90.

    Article  CAS  PubMed  Google Scholar 

  32. Rockenbach II, Jungfer E, Ritter C, Santiago-Schubel B, Thiele B, Fett R, Galensa R. Characterization of flavan-3-ols in seeds of grape pomace by CE, HPLC-DAD-MSn and LC-ESI-FTICR-MS. Food Res Int. 2012;48(2):848–55.

    Article  CAS  Google Scholar 

  33. Zhang ZR, Al Zaharna M, Wong MM, Chiu SK, Cheung HY. Taxifolin enhances andrographolide-induced mitotic arrest and apoptosis in human prostate cancer cells via spindle assembly checkpoint activation. PLoS ONE. 2013;8(1):e54577.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Watjen W, Michels G, Steffan B, Niering P, Chovolou Y, Kampkotter A, Tran-Thi QH, Proksch P, Kahl R. Low concentrations of flavonoids are protective in rat H4IIE cells whereas high concentrations cause DNA damage and apoptosis. J Nutr. 2005;135(3):525–31.

    PubMed  Google Scholar 

  35. He Z, Xia W. Analysis of phenolic compounds in Chinese olive (Canarium album L.) fruit by RPHPLC-DAD-ESI-MS. Food Chem. 2007;105(3):1307–11.

    Article  CAS  Google Scholar 

  36. Perestrelo R, Lu Y, Santos SAO, Silvestre AJD, Neto CP, Câmara JS, Rocha SM. Phenolic profile of Sercial and Tinta Negra Vitis vinifera L. grape skins by HPLC–DAD–ESI-MSn: novel phenolic compounds in Vitis vinifera L. grape. Food Chem. 2012;135(1):94–104.

    Article  CAS  Google Scholar 

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Authors’ contributions

AS and AR designed experiments, conducted the experimental work, and prepared the manuscript; IHS, VvG and RS also helped AS to conduct the experimental work and analyze the data. HS collected, identified and prepared plant materials. NK and MU organized, discussed and supervised the project and contributed to manuscript writing. All authors read and approved the final manuscript.

Acknowledgements

The authors would like to thank Ms. Anja Müller for the technical support and Dr. Hartwig Schepker for providing the Rhododendron material and their complete botanical information.

Competing interests

The authors declare that they have no competing interests.

Availability of data and materials

A supplement titled “Additional file 1.docx” has been uploaded along with the manuscript, which includes the chromatogram of the different parts of R. ambiguum, the structures and fragmentation patterns of all compounds identified in this study.

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The authors hereby consent to the publication of the manuscript in the journal BMC Research Notes.

Funding

The authors would like to thank Rhododendron Park and Jacobs University for their financial support.

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Authors and Affiliations

  1. Department of Life Science & Chemistry, Jacobs University Bremen, Campus Ring 1, 28759, Bremen, Germany

    Abhinandan Shrestha, Ahmed Rezk, Inamullah Hakeem Said, Victoria von Glasenapp, Rachelle Smith, Matthias S. Ullrich & Nikolai Kuhnert

  2. Stiftung Bremer Rhododendronpark, Deliusweg 40, 28359, Bremen, Germany

    Hartwig Schepker

Authors
  1. Abhinandan Shrestha
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  2. Ahmed Rezk
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  3. Inamullah Hakeem Said
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  4. Victoria von Glasenapp
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  5. Rachelle Smith
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  6. Matthias S. Ullrich
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  7. Hartwig Schepker
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  8. Nikolai Kuhnert
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Corresponding author

Correspondence to Nikolai Kuhnert.

Additional file

13104_2017_2601_MOESM1_ESM.docx

Additional file 1: Figure A1. Total Ion Chromatogram of Rhododendron ambiguum first year leaves, second year leaves, flowers and fruits generated by LC-MSn in negative mode. Figure A2. Structures of all identified compounds in Rhododendron ambiguum and Rhododendron cinnabarinum. Table A1. Fragmentation pattern of the identified compounds in Rhododendron ambiguum and Rhododendron cinnabarinum.

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Shrestha, A., Rezk, A., Said, I.H. et al. Comparison of the polyphenolic profile and antibacterial activity of the leaves, fruits and flowers of Rhododendron ambiguum and Rhododendron cinnabarinum . BMC Res Notes 10, 297 (2017). https://doi.org/10.1186/s13104-017-2601-1

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BMC Research Notes

ISSN: 1756-0500

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