J Food Biochem. 2020;44:e13107. wileyonlinelibrary.com/journal/jfbc | 1 of 11 https://doi.org/10.1111/jfbc.13107
© 2019 Wiley Periodicals, Inc. Received: 15 August 2019
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Revised: 18 October 2019|
Accepted: 11 November 2019DOI: 10.1111/jfbc.13107 F U L L A R T I C L E
Chemical profile, antiproliferative, antioxidant, and enzyme
inhibition activities and docking studies of Cymbopogon
schoenanthus (L.) Spreng. and Cymbopogon nervatus (Hochst.)
Chiov. from Sudan
Sakina Yagi
1| Atif B. A. Mohammed
1| Tzvetomira Tzanova
2| Hervé Schohn
3|
Haider Abdelgadir
4| Azzurra Stefanucci
5| Adriano Mollica
5| Gökhan Zengin
61Faculty of Science, Department of Botany, University of Khartoum, Khartoum, Sudan 2Department of Biosis, Université de Lorraine, CNRS, L2CM, Nancy, France 3Department of Biosis, Université de Lorraine, CNRS, CRAN, Nancy, France 4Faculty of Science, Department of Biology, Albaha University, Saudi Arabia
5Department of Pharmacy, University “G. d’Annunzio” of Chieti-Pescara, Chieti, Italy 6Science Faculty, Department of Biology, Selcuk University, Konya, Turkey Correspondence
Gökhan Zengin, Science Faculty, Department of Biology, Selcuk University, Campus, Konya, Turkey.
Email: gokhanzengin@selcuk.edu.tr
Abstract
Essential oils from the inflorescence of Cymbopogon schoenanthus and C. nervatus growing in Northern Sudan were examined for their chemical composition, antipro-liferative activity against human breast carcinoma and human colon adenocarcinoma cell lines, antioxidant activity (phosphomolybdenum, antiradical, reducing power, and ferrous chelating), and enzyme inhibition activity against acetylcholinesterase bu-tyrylcholinesterase, tyrosinase, α-glucosidase, and α-amylase. In silico study on the inhibition of tyrosinase and α-amylase was also performed. Piperitone (59.1%) and isomers of para-menthadienols (35.3%) were the main compounds in C.
schoenan-thus and C. nervatus oils, respectively. Oil from C. nervatus possessed higher
anti-oxidant activity than that from C. schoenanthus except for its metal chelating ability. Both oils showed high antiproliferative activity. In silico study showed that trans-p-mentha-2,8-dien-1-ol and piperitone (both isomers) revealed the best docking scores for α-amylase and tyrosinase, respectively. In conclusion, oils from these two
Cymbopogon species could be new natural agents with functional properties for food,
cosmetics, and pharmaceutical industries.
Practical applications
Recently, there is a growing tendency to replace synthetic oils by natural ones in the cosmetic, food, and pharmaceutical products. In this context, we investigated the chemical characterization and biological activities of two Cymbopogon species essen-tial oils (C. schoenanthus (L.) Spreng. and C. nervatus). Antioxidant capacity, enzyme inhibition, and antiproliferative effects were tested for biological activities. Chemical characterization was identified by GC-MS. Based on our findings, the Cymbopogon species may be utilized as sources of natural bioactive agents in food industries.
K E Y W O R D S
1 | INTRODUCTION
From ancient times to today, essential oils have wide applications in food, traditional medicine, cosmetics, and pharmaceutical industries. The essential oil market is expected to reach $11.5 billion in 2022 (Allied, 2016). More than 3,000 plants are known to produce essen-tial oils of which approximately 300 are of commercial importance (Palazzolo, Laudicina, & Germanà, 2013). About 65% of the world's production of essential oils came from developing countries. Studies on essential oils from aromatic plants showed that they exert a broad spectrum of bioactivity owing to the presence of volatile molecules such as terpenes and phenol-derived aromatic and aliphatic compo-nents (Adefegha, Olasehinde, & Oboh, 2017; Lawal & Ogunwande, 2013; Oboh, Olasehinde, & Ademosun, 2017).
Aromatic plants of Africa are currently receiving great attention due to their biological and chemical diversities. Many of these plants are widespread throughout the world, and important in African tra-ditional medicine, being used for the treatment of diseases such as malaria, hepatitis, cancer, inflammation, and infections by fungi, bac-teria, and viruses (Bursal et al., 2019; Gülçin, Tel, Gören, Taslimi, & Alwasel, 2019; Lawal & Ogunwande, 2013; Tohma et al., 2016).
The genus Cymbopogon (family Poaceae) is widely spread throughout the world and comprised about 144 species distributed in the tropical and subtropical Africa, Asia, and Australia (Khanuja et al., 2005). Of the nine Cymbopogon species present in Sudan, two,
C. schoenanthus (L.) Spreng. (commonly known as maharaib) and C. nervatus (Hochst.) Chiov. (commonly known as nal) are of economic
importance (Modawi, Magar, Satti, & Duprey, 1984). In Sudanese tra-ditional medicine, the aerial part of C. schoenanthus is used as stom-achache (El Ghazali, El-Tohami, & El-Egami, 1994), antispasmodic (Suleiman, 2015), and to treat diabetes (Issa et al., 2018). Inflorescence of C. nervatus is used for the treatment of kidney pains and urethritis (El-Kamali, Hamza, & El-Amir, 2005) and the leaves against indiges-tion, as carminative and tonic (El-Kamali & El-Khalifa, 1999).
Previous pharmacological studies showed that the essential oil of C. schoenanthus possessed insecticidal (Ketoh, Koumaglo, Glitho, & Huignard, 2006), antioxidant (Khadri et al., 2008), antiacetylcho-linesterase (AChE) (Khadri et al., 2008), anthelmintic (Katiki, Chagas, Bizzo, Ferreira, & Amarante, 2011), antibacterial (Hashim, Almasaudi, Azhar, Al Jaouni, & Harakeh, 2017), antifungal, and anti-inflamma-tory (Norbert et al., 2014) activities. The essential oil of C. nervatus inflorescence was shown to exhibit antibacterial (El-Kamali et al., 2005) and spasmolytic (Omar et al., 2016) activities.
Recently, there is a growing tendency to replace synthetic oils by natural ones in the cosmetic, food, and pharmaceutical products (Džamić & Matejić, 2017). The objective of this study was aimed at the investigation of the chemical profile of essential oils from C.
schoe-nanthus and C. nervatus inflorescence collected from Northern Sudan
and to evaluate their antiproliferative, antioxidant, and enzyme inhi-bition activities. Antiproliferative activity was evaluated against three cell lines established from human breast carcinoma samples (MCF7) and human colon adenocarcinoma samples (HT29 and HCT116) as well as Vero cells (established from African green monkey kidney and
represented an untransformed cell line). Antioxidant activity included scavenging of free radicals, reduction potential, phosphomolybdenum, and chelating ability. Enzyme inhibition properties were evaluated on enzymes related to neurodegenerative ailments (AChE and buty-rylcholinesterase [BuChE]), diabetes and obesity (α-glucosidase and α-amylase), and skin hyperpigmentation (tyrosinase).
2 | MATERIALS AND METHODS
2.1 | Plant materials
Inflorescence of C. schoenanthus and C. nervatus was collected in January 2014 from Shendi, a city located in North Sudan (longitude: 33° 26ʹ E; latitude 16° 41ʹ N). Plants were taxonomically identified and voucher specimens (No. 2014/1CS for C. schoenanthus and No. 2014/1CN for
C. nervatus) have been deposited in the Botany Department Herbarium,
Faculty of Science, University of Khartoum, Sudan.
2.2 | Extraction of essential oils
Essential oils from all the plant species (1 kg) were extracted by hydr-odistillation using a Clevenger-type apparatus for 4 hr. The extracted oils were dried over anhydrous sodium sulphate and stored at 4°C, in amber-colored bottles, before use.
2.3 | Chemical analysis of the essential oils
Agilent 5975 GC-MSD system coupled with an Agilent 7890A GC (Agilent Technologies Inc., Santa Clara, CA) was used to determine the chemical profile of the obtained essential oils (in n-hexane). HP-INNOWax FSC column (60 m × 0.25 mm, 0.25 μm film thickness) was used as a column and all analytical parameters have been reported in our previous paper (Zengin, Sarıkürkçü, Aktümsek, & Ceylan, 2016). We calculated the retention index of each component by a homol-ogous series of n-alkanes (C8-C30), under the same experimental conditions. In order to further provide identifications, we compared unknown compound spectra with libraries such as NIST 05 and Wiley 8th version. To quantify essential oil components, we used the relative area method, which reflects in a percentage relation of the area corresponding to each component with regards to the total area of the obtained chromatogram.
2.4 | Antiproliferative activity
Antiproliferative activity of oils against human breast carcinoma (MCF7), human colon adenocarcinoma (HT29 and HCT116), and nor- mal Vero cell lines was evaluated using the 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) procedure as described by Mosmann (1983).
2.4.1 | Cell culture
HCT116 and HT29 were cultivated in Dulbecco's minimum essen-tial medium (DMEM,) supplemented with 10% fetal calf serum, 1% penicillin/streptomycin, and 2 mM L-glutamine. MCF7 and Vero cells were grown in RPMI medium with the same additives.2.4.2 | MTT procedure
In brief, cells were seeded in 96-well plate at 10,000 cells/well for HT29, MCF-7, and Vero cells and at 5,000 cells/well for HCT116 cells. About 24 hr after seeding, 100 μl of medium containing in-creasing concentrations (range from 0.5 to 400.0 μg/ml) of each es-sential oil was added to each well for 72 hr at 37°C. After incubation, the medium was discarded and 100 μl/well of MTT solution (0.5 mg/ ml) was added and incubated for 2 hr. Water-insoluble formazan blue crystals were finally dissolved in DMSO at 200 μl/well. Each plate was read at 570 nm. The viability was evaluated based on a compari-son with untreated cells. IC50 was calculated using GraphPad Prism(GraphPad Software, La Jolla, CA, USA).
2.5 | Antioxidant activity
The antioxidant potential of the oils was evaluated by phosphomo-lybdenum, antiradical (DPPH and ABTS), reducing power (FRAP and CUPRAC), and ferrous chelating assays as described by Grochowski et al. (2017). Trolox equivalents were used for the expression of anti-oxidant activities. EDTA was employed as a reference compound for the metal chelating assay.
2.6 | Enzyme inhibition activity
The key enzymes’ inhibition activity of extracts against ACh, BuCh, tyrosinase, α-glucosidase, and α-amylase were measured using the protocols by Grochowski et al. (2017)
2.7 | Molecular modeling
2.7.1 | Enzymes preparation
The major components of essential oil of Cymbopogon proximus and C. nervatus have taken into consideration for in silico evalua-tion studies, in order to elucidate the binding mode to the enzymatic pocket of tyrosinase and α-amylase. The crystal structures of ty-rosinase (pdb:2y9x) (Ismaya et al., 2011) in complex with tropolone, α-amylase (pdb:4gqr) (Williams, Li, Withers, & Brayer, 2012) in complex with myricetin have been downloaded from the PDB da-tabase available online (Berman et al., 2000). The enzymes have been prepared for docking by PrepWizard (Sastry, Adzhigirey, Day,
Annabhimoju, & Sherman, 2013) tool embedded in Maestro 2015 (Shroedinger, 2015). This software was used to neutralize the macro-molecules at pH 7.4 by PROPKA (Shelley et al., 2007) to convert the seleno-cysteines and seleno-methionines, if present, respectively, to cysteines and methionines and to fix other errors present in the raw structure of the enzymes. All the missing fragments and other errors present in the crystal structures were automatically solved and the hydrogens were added and minimized following the general method previously reported by our previous papers (Mahomoodally et al., 2018; Mocan, Zengin, Crisan, & Mollica, 2016; Mocan et al., 2017) with the use of OPLS3 force field (Harder et al., 2016).
2.7.2 | Ligands preparation
The selected components of the essential oils submitted to the docking are: α-pinene (2), camphene (4), δ-2-carene (5), isocin-eole (10), limonene (12), 1,8-cinisocin-eole (13), p-cymene (16), terpi-nolene (17), α,p-dimethylstyrene (20), camphor (23), cis-dihydro α-terpineol (25), trans-p-menth-2-en-1-ol (26), 1-terpineol (27), fen-chol (29), β-elemene (30), terpinen-4-ol (32), β-caryophyllene (33), trans-p-mentha-2,8-dien-1-ol (36), cis-p-mentha-2,8-dien-1-ol (37), α-terpineol (39), borneol (40), piperitone (44(+) and 44(−)), carvone (45), trans-p-mentha-7,8-dien-2-ol (49), cuparene (50), trans-carveol (51), p-cymen-8-ol (53), cis-carveol (54), cis-p-mentha-7,8-dien-2-ol (55), caryophyllene oxide (56), elemon (57), (E)-methyl cinnamate (58), γ-eudesmol (59), α-eudesmol (61) and β-eudesmol (62). The compounds have been downloaded from Zinc database of drawn by ChemBioDraw 14.0 software and prepared by LigPrep module em-bedded in Maestro 10.2 (Schrödinger, 2012).
2.7.3 | Molecular docking
The docking experiments have been carried out on tyrosinase by GOLD (Verdonk, Cole, Hartshorn, Murray, & Taylor, 2003) and on α-amylase by the software Glide by performing a Standard Precision docking and resubmitting the so obtained poses to docking proce-dure employing the eXtra Precision method (Friesner et al., 2006). The docking area was delimited automatically by the software, gen-erating a docking grid centered at the crystallographic inhibitor and extended in a radius of 10 Angstroms around the ligand center for Gold, in case of Glide the grid is a box of 20×20×20 Angstroms. The docking scores obtained for the essential oil components tyrosinase and α-amylase are reported in Table 1.
2.8 | Statistical analysis
The results of antioxidant and enzyme inhibitory activities are given as mean ± standard deviation (SD). The results were statistically evaluated using the Student's t test (α = .05). Statistical analysis was carried out using SPSS v. 14.0 program.
3 | RESULTS AND DISCUSSION
3.1 | Chemical profile of essential oils from
C. schoenanthus and C. nervatus
Dry inflorescence of C. schoenanthus yielded 1.0% (w/w) of yel-low-colored oil, while 0.8% (w/w) of light yellow-colored oil was obtained from C. nervatus. About 29 compounds, representing 99.4% of the total oil, were identified in C. schoenanthus with high dominance of oxygenated monoterpenes (62.1%) followed by
oxygenated sesquiterpenes (21.6%) and hydrogenated monoter-penes (11.6%), respectively (Table 2). A total of 42 compounds, corresponding to 90.6% of the total oil, were recognized from the oil of C. nervatus where the majority of constituents belonged to oxygenated monoterpenes (66.1%) followed by hydrogen-ated monoterpenes (22.2%). As previously reported by Omar et al. (2016) sesquiterpenes were scare in C. nervatus constituting only 1%. Piperitone (59.1%) represented the major constituent in C. schoenanthus, while limonene (13.7%) and isomers of para-menthadienols including trans-p-mentha-2,8-dien-1-ol (13.6%),
No Compounds α-Amylase docking score Tyrosinase GoldScore
2 α-pinene (+) and α-pinene (−) −3.24 and −3.02 30.1 and 28.8
4 Camphene n.a. 32.8 5 δ-2-carene −3.45 33.3 10 Isocineole −3.67 36.7 12 Limonene −3.10 36.2 13 1,8-cineole −2.6 28.3 16 p-cymene −2.93 42.1 17 terpinolene −3.10 39.1 23 Camphor n.a. 25.1 25 Cis-dihydro a-terpineol −5.01 34.7 26 Trans-p-menth2-en-1-ol −4.47 40.6 27 1-terpineol −3.50 38.8 29 Fenchol −3.29 26.5 30 β-elemene n.a. 41.4 32 Terpinen-4-ol −5.13 37.6 33 β-caryophyllene −3.02 37.7 36 Trans-p-mentha-2,8-dien-1-ol −4.66 47.8 37 Cis-p-mentha-2,8-dien-1-ol −5.20 40.6 39 α-terpineol −4.75 39.4 40 Borneol n.a. 19.9
44 Piperitone (+) and (−) −5.20 and −5.30 37.1 and 38.9
45 carvone −4.7 36.9 49 Trans-p-mentha-1(7),8-dien-2-ol −4.43 37.0 50 Cuparene −3.38 32.4 51 Trans-carveol −5.24 38.4 53 p-cymen-8-ol −5.02 41.5 54 Cis-carveol n.a. 37.6 55 Cis-p-mentha-1(7),9-dien-2-ol −4.52 36.5 56 Caryophyllene oxide −4.42 31.3 57 Elemol −3.73 40.0 58 (E)-methyl cinnamate n.a. 38.1 59 γ-eudesmol −4.67 33.5 61 α-eudesmol −3.58 33.3 62 β-eudesmol −4.00 33.4
Note: Bold numbers represent the best values.
TA B L E 1 Docking scores and fitness
GoldScore of selected compounds docked on α-amylase and tyrosinase
TA B L E 2 Chemical composition of essential oils from inflorescences of Cymbopogon proximus and C. nervatus
No. Compounds RRIa
Percentage occurrence (%) C. proximus C. nervatus 1 Tricyclene 1,009 – trb 2 α-Pinene 1,023 – 0.6 3 α-Fenchene 1,057 – 0.1 4 Camphene 1,068 – 0.6 5 δ-2-Carene 1,132 9.1 tr 6 δ-3-Carene 1,157 – 0.1 7 Myrcene 1,165 – tr 8 α-Phellandrene 1,168 0.1 tr 9 Pseudolimonene 1,173 – tr 10 Isocineole 1,179 – 4.5 11 α-Terpinene 1,183 – 0.2 12 Limonene 1,201 1.9 13.7 13 1,8-Cineole 1,211 – 3.2 14 p-Mentha-1,3,6-triene 1,218 – 0.2 15 γ-Terpinene 1,249 – 0.3 16 p-Cymene 1,276 0.4 3.1 17 Terpinolene 1,286 – 2.6 18 Isoterpinolene 1,291 – 0.1 19 α-Fenchone 1,408 0.1 – 20 α,p-dimethylstyrene 1,447 0.1 0.6 21 Longipinene 1,479 – 0.1 22 Longicyclene 1,512 – 0.1 23 Camphor 1,535 – 0.3 24 Sativene 1,539 – tr 25 cis-Dihydro α-terpineol 1,567 – 0.3 26 trans-p-Menth-2-en-1-ol 1,568 0.4 – 27 1-Terpineol 1,580 – 3.1 28 Longifolene 1,585 – 0.8 29 Fenchol 1,592 1.4 30 β-Elemene 1,601 1.2 – 31 Calarene (=β-Gurjuene) 1,606 0.1 – 32 Terpinen-4-ol 1,612 – 0.5 33 β-Caryophyllene 1,614 0.6 – 34 trans-Dihydrocarvone 1,626 – 0.1 35 cis-p-Menth-2-en-1-ol 1,633 0.5 – 36 trans-p-Mentha-2,8-dien-1-ol 1,636 – 13.6 37 cis-p-Mentha-2,8-dien-1-ol 1,677 tr 7.9 38 trans-Piperitol 1,689 0.1 – 39 α-Terpineol 1,706 1.1 3.2 40 Borneol 1,715 – 0.9 41 Chamigrene 1,723 0.1 – 42 β-Selinene 1,743 0.4 – 43 α-Selinene 1,747 0.2 – 44 Piperitone 1,753 59.1 – (Continues)
trans-p-mentha-1(7),8-dien-2-ol (11.1%), and cis-p-mentha-1(7),8-dien-2-ol (10.6%) formed the major compounds in C. nervatus. Generally, these results were in agreement with previous data obtained from plants growing in Sudan ((Elhassan, Eltayeb, & Khalafalla, 2016; Yagi, Babiker, Tzanova, & Schohn, 2016), for C.
schoenanthus and (Omar et al., 2016) for C. nervatus) or from other
countries (Hakkim, Al-Balushi, & Achankunju, 2016; Menut et al., 2000; Selim, 2011) for C. schoenanthus). The only difference was mainly in the proportion of compounds that could be attributed to different biotic and abiotic factors.
3.2 | Antiproliferative activity
The results of the antiproliferative activity of essential oils from C.
sch-oenanthus and C. nervatus inflorescence are depicted in Table 3. The two
extracted oils were not toxic to normal Vero cell line (IC50 > 100 µg/ml).
Although both oils showed different chemical profiles they exhibited
high and comparable antiproliferative activity against the three tested cancer cell lines with IC50 values comprised between 4.3 and 5.9 µg/ ml. These results were remarkably different from those reported by Yagi et al. (2016) who found that the antiproliferative activity of
C. schoenanthus aerial parts against these cell lines ranged between
19.16 and 38.43 µg/ml in the same cell lines. This notable variation in
No. Compounds RRIa
Percentage occurrence (%) C. proximus C. nervatus 45 Carvone 1,757 – 2.9 46 δ-Cadinene 1,773 0.1 – 47 γ-Cadinene 1,779 0.1 – 48 p-Methyl acetophenone 1,800 – 0.3 49 trans-p-mentha-1(7),8-dien-2-ol 1,809 – 11.1 50 Cuparene 1,846 0.5 – 51 trans-Carveol 1,846 – 2.2 52 Geraniol 1,852 0.2 – 53 p-Cymen-8-ol 1,861 0.6 0.9 54 cis-Carveol 1,877 – 0.5 55 cis-p-mentha-1(7),8-dien-2-ol 1,900 – 10.6 56 Caryophyllene oxide 2,017 0.7 – 57 Elemol 2,095 10.9 – 58 (E)-Methyl cinnamate 2,104 0.9 – 59 γ-Eudesmol 2,187 2.1 – 60 Eugenol 2,188 – 0.3 61 α-Eudesmol 2,246 3.0 – 62 β-Eudesmol 2,256 4.9 – 63 Acetyl eugenol 2,276 – 0.5 Monoterpenes hydrocarbons 11.6 22.2 Oxygenated monoterpenes 62.1 66.1 Sesquiterpenes hydrocarbons 3.3 1 Oxygenated sesquiterpenes 21.6 0 Others 0.8 1.3 Total 99.4 90.6 Extraction yield 1.0% 0.8%
aRelative retention indices calculated against n-alkanes. bTrace (<0.1%).
TA B L E 2 (Continued)
TA B L E 3 Antiproliferative activity of essential oils from
inflorescences of Cymbopogon proximus and C. nervatus
Name IC50 (µg/ml) Vero HT29 HCT116 MCF7 C. proximus 4.3 ± 0.5 4.4 ± 0.5 5.8 ± 0.8 >100 C. nervatus 4.4 ± 0.9 4.3 ± 0.5 5.9 ± 0.9 >100 Note: Results were obtained from the quadruplicate determination of two independent experiments (n = 8 using increasing concentrations (ranged from 0.5 to 400.0 μg/ml) of each essential oil.
antiproliferative activity could be attributed to the composition and concentrations of essential oils which are mainly influenced by several factors including plant parts, stage of vegetative cycle, seasonal varia-tion, geographical source, climatic and soil conditions, (Callan, Johnson, Westcott, & Welty, 2007). In fact, in this study, oil was extracted from a sample of C. schoenanthus inflorescences harvested from North Sudan, while that studied by (Yagi et al., 2016) was obtained from C. schoenan-thus leafy stems growing in Western Sudan.
3.3 | Antioxidant activity
Five complementary assays were performed for the evaluation of the antioxidant capacity of the essential oils of these two Cymbopogon spe-cies and results are presented in Table 4. The free radical scavenging capacity of the oils was measured using DPPH and ABTS radical cation. Oil from C. nervatus areal parts displayed significantly (p < .05) higher DPPH (15.70 ± 0.29 mg TEs/g oil) and ABTS (68.32 ± 1.99 mg TEs/g oil) scavenging activity than that from C. schoenanthus (14.61 ± 0.37 and 63.93 ± 1.16 mg TEs/g oil, respectively) with particularly higher activity observed from the ABTS assay. Ferric and cupric ion reducing the power of the oils were determined by CUPRAC and FRAP assays. Oil of C. nervatus showed significantly (p < .05) higher reducing power activity (144.22 ± 0.33CUPRAC and 131.48 ± 2.98FRAP mg TEs/g oil) than
that of C. schoenanthus (133.20 ± 2.36CUPRAC and101.95 ± 1.12FRAP mg TEs/g oil). The metal chelating ability of the two oils was determined by their capacity to capture Fe2+ and results showed that oil of C.
sch-oenanthus (31.40 ± 1.22 mgEDTAEs/g oil) exerted significant (p < .05)
higher metal chelating ability than that of C. nervatus (25.00 ± 3.27 mgEDTAEs/g oil). The total antioxidant activity determined from phosphomolybdenum assay revealed that oil of C. nervatus was sig-nificantly (p < .05) more active (2 times) than that of C. schoenanthus
(26.60 ± 1.13 mmolTEs/g oil). Thus, it was clear that, oil from C.
ner-vatus possessed higher antioxidant activity than that from C. schoe-nanthus except for its metal chelating ability where C. schoenanthus oil
revealed better capacity. Previous studies on the antioxidant activity of extracted oils from these two species were only determined by the DPPH and/or β-carotene–linoleic acid bleaching methods. Khadri et al. (2008) showed that the highest antioxidant activity of C. schoenanthus oil grown in Tunisia was obtained from extracted oil prepared from fresh leaves collected in the desert region and dried roots collected from the mountain using β-carotene–linoleic acid bleaching and DPPH methods, respectively. (Omar et al., 2016) reported that oil from C.
ner-vatus inflorescence grown in Sudan showed moderate DPPH radicals
activity.
3.4 | Enzyme inhibition activity
AChE is a key hydrolytic enzyme that is responsible for the reduc-tion of acetylcholine (ACh) levels in the hippocampus and cortex of the brain and inhibition of this enzyme is considered as an appropri-ate strategy to treat Alzheimer disease (Zengin et al., 2015). In this study, oils from C. schoenanthus and C. nervatus showed compara-ble AChE (1.24 ± 0.09 and 1.26 ± 0.05 mg GALAEs/g, respectively) and BChE (1.21 ± 0.03 and 1.14 ± 0.05 mg GALAEs/g, respectively) inhibition ability (Table 5). Previous study on anti-AChE activity of these two Cymbopogon species was only carried for oil and extracts of C. schoenanthus growing in Tunisia where the best AChE inhibi-tory activity was exhibited by the leaves oil and ethyl acetate and methanol extracts of the plants collected from the mountainous region (Khadri et al., 2010, 2008). Tyrosinase is a vital enzyme in many skin conditions like hyperpigmentation and also in dopamine toxicity in Parkinson's disease and inhibition of this enzyme could
TA B L E 4 Antioxidant activity of essential oils from inflorescences of Cymbopogon proximus and C. nervatus
Essential oils DPPH (mgTEs/g)* ABTS (mgTEs/g)* CUPRAC (mgTEs/g)* FRAP (mgTEs/g)*
Metal Chelating (mgEDTAEs/g)** Phosphomolybdenum (mmolTEs/g)* C. proximus 14.61 ± 0.37b 63.93 ± 1.16b 133.20 ± 2.36b 101.95 ± 1.12b 31.40 ± 1.22a 26.60 ± 1.13b C. nervatus 15.70 ± 0.29a 68.32 ± 1.99a 144.22 ± 0.33a 131.48 ± 2.98a 25.00 ± 3.27b 53.06 ± 3.62a Note: Different superscript letters (a and b) in the same column indicate a significant difference (p < .05); na, not active. *TEs, Trolox equivalents. **EDTAEs, disodium edetate equivalents.
TA B L E 5 Enzyme inhibitory activity of essential oils from inflorescences of Cymbopogon proximus and C. nervatus
Essential oils Acetylcholinesterase (mg GALAEs/g)* Butyrylcholinesterase (mg GALAEs/g)* Tyrosinase
(mg KAE/g)** α-amylase (mg ACAEs/g)*** α-glucosidase (mgACAEs/g)***
C. proximus 1.24 ± 0.09a 1.21 ± 0.03a 68.80 ± 3.15a 55.22 ± 1.16a na C. nervatus 1.26 ± 0.05a 1.14 ± 0.05a 37.09 ± 0.16b 47.85 ± 1.02b na Note: Different superscript letters (a and b) in the same column indicate a significant difference (p < .05). *GALAEs, galanthamine equivalents. **KAEs, kojic acid equivalents. ***ACEs, acarbose equivalents.
be beneficial for the treatment of such diseases (Erdogan Orhan & Tareq Hassan Khan, 2014). Both oils displayed good inhibitory activity against tyrosinase but the oil from C. schoenanthus exhib- ited significantly (p < .05) higher (1.9-fold) tyrosinase inhibition abil-ity than that from C. nervatus (37.09 ± 0.16 mg KAE/g) (Table 4). The inhibition of α-amylase and α-glucosidase, key enzymes in hy-drolysis of starch and oligosaccharide, control the blood glucose level and thus counted as an important strategy in the manage-ment of diabetes mellitus (Mnafgui et al., 2016). Inhibitory activity of the two oils against these two digestive enzymes is presented in Table 4. Oil of C. schoenanthus (55.22 ± 1.16 mg ACAEs/g) re-vealed significantly (p < .05) higher α-amylase inhibition activity than that of C. nervatus (47.85 ± 1.02 mg ACAEs/g). However, both oils did not exert any α-glucosidase inhibition activity. To the best of our knowledge, this is the first report on the BChE, tyrosinase,
α-amylase, and α-glucosidase inhibition activities of these two
Cymbopogon species.
Essential oils are complex mixtures and their bioactivity is not clearly understood. In some cases, isolated constituents were tested such as thymol, thymol acetate, and carvacrol. These molecules ex-hibited anticholinesterase activity higher than the whole oil, sug-gesting that the enzyme inhibition activity of the oil is mainly due to the presence of a specific component in a significant proportion (Jukic, Politeo, Maksimovic, Milos, & Milos, 2007; Silva et al., 2019). In this context, it is likely that the major compounds, piperitone (59.1%) in C. schoenanthus oil and isomers of para-menthadienols (35.3%) in C. nervatus, could be largely responsible for the observed antiproliferative, antioxidant, and enzyme inhibition activities of these two oils. Furthermore, it was demonstrated that limonene, which represented 13.7% of C. nervatus oil, exerts both antioxidant
F I G U R E 1 Docking dose depiction of compound 36 docked to tyrosinase (a and b); docking pose of compound 44 docked to α-amylase (c
(Junior et al., 2009) and anti-proliferative (Bayala et al., 2014; Manuele, Ferraro, & Anesini, 2008) activities. Furthermore, it was hypothesized that the main components are not necessarily respon-sible for the higher share of the total activity of the extracted oil and constituents with low abundance should be considered too (Saidana et al., 2008). For example, terpinen-4-ol, which was found in a low proportion (0.5%) in C. nervatus oil, has been described previously to possess anticancer properties and induce apoptosis (Döll-Boscardin et al., 2012). Besides, α-terpineol, constituting 1.1% and 3.2% of C.
schoenanthus and C. nervatus oils, was found to possess high
radi- cal-scavenging activity (Miguel, 2010). Moreover, some studies sug-gested that the action of essential oils cannot be assigned to only one compound, notably the major component. Diverse molecules of oils may act synergistically leading to an overall activity that results from the interaction among several compounds (Chouhan, Sharma, & Guleria, 2017). This was demonstrated with components like cam-phor, 1,8-cineole, α-pinene. geraniol, linalool, and γ-terpinene. These molecules were not as potent as the total oil to inhibit cholinester-ase, suggesting the possibility of a synergistic action of two or more compounds which mutually regulate each other (Orhan, Kartal, Kan, & Şener, 2008; Perry, Houghton, Theobald, Jenner, & Perry, 2000). For example, Miguel et al. (2018) showed that 1,8-cineole, which constituted 3.2% of C. nervatus oil, has a low effect on THP-1 human leukaemia cells. However, the antiproliferative activity increased in the presence of p-cymene (representing 3.1% of C. nervatus oil) sup-porting the concept of a synergistic effect of both molecules.
3.5 | Molecular modeling
We have examined by in silico molecular docking, the most abundant substances present in both extracts object of this work. The values of the docking scores (from maestro) for α-amylase reveal that the best interactions with the enzyme has been found for the com-pounds 25, 32, 44, 51, 53; on the other hand for the tyrosinase, the best poses obtained by GOLD are for compounds 16, 26, 30, 36, 37,
53, 57. For some of the analyzed substances, the biological activity
was already reported in the literature, for example, for 32 (Azhar Ul et al., 2006) and 58 (Iscan, Kirimer, Kurkcuoglu, Baser, & Demirci, 2002). Based on our results, also compound 36 could be taken into consideration for antityrosinase activity and compound 44 (both iso-mers) as a α-amylase inhibitor.
Compound 36, namely trans-p-Mentha-2,8-dien-1-ol, in its best docking pose was found to be able to establish to the enzymatic pocket of tyrosinase several van der Waals interactions and two da-tive bonds to the Copper atoms of the enzyme. In particular, this latest feature is the discriminant factor that should allow this com-pound to interact better than the others to the enzyme (for the best pose depiction see Figure 1a,b).
Compound 44, namely piperitone, was the best performer for the ss could be present in the extract in both (+) and (−) isomers, thus the two compounds have been tested separately. The results of the docking, considering the docking scores obtained, have shown that
both isomers have the same affinity for the enzyme. In Figure 1b, the compound represented is the isomer (−). Both compounds are able to interact in the cavity of the α-amylase by forming 1 H-bond with Gln63 and several lipophilic interactions with the hydrophobic groups of Leu162, Leu165, Tyr62, and Trp58 (Figure 1c,d). However, in vitro studies are required to provide a confirmation and detailed understanding of the interactions between the enzyme and these compounds.
4 | CONCLUSIONS
In this study, oxygenated monoterpenes represented the main com-pounds of extracted oil from dried inflorescences of C. schoenanthus and C. nervatus. Both oils exhibited marked antiproliferative activity against cancer cell lines as well as considerable antioxidant potential acting by different mechanisms. Moreover, the oils indicated a high capacity to inhibit some key enzymes that are associated with major health problems including Alzheimer's disease, skin disorders, and diabetes mellitus. To obtain better valorization of possible therapeu-tic application of these oils, further in vivo studies should be car-ried out to evaluate their bioactive properties and to explore their mechanism of action.ACKNOWLEDGMENTS
SY and ABAM would like to express their gratitude to Mr. Ahmed Saeed (Institution of Aromatic and Medicinal Plants) for his generous assistance during the study.
CONFLIC T OF INTEREST
There are no conflicts of interest to declare.
ORCID
Sakina Yagi https://orcid.org/0000-0002-9600-3526
Hervé Schohn https://orcid.org/0000-0002-3651-2636
Azzurra Stefanucci https://orcid.org/0000-0001-7525-2913
Adriano Mollica https://orcid.org/0000-0002-7242-4860
Gökhan Zengin https://orcid.org/0000-0001-6548-7823 REFERENCES
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How to cite this article: Yagi S, Mohammed ABA, Tzanova T,
et al. Chemical profile, antiproliferative, antioxidant, and enzyme inhibition activities and docking studies of
Cymbopogon schoenanthus (L.) Spreng. and Cymbopogon nervatus (Hochst.) Chiov. from Sudan. J Food Biochem.
