Jurnal Kimia Sains danAplikasi 24 (5) (2021):152-160 152

Jurnal Kimia Sains danAplikasi 24 (5) (2021):152-160ISSN:1410-8917

Jurnal Kimia-Sains &Aplikasi

e-ISSN:2597-9914

Jurnal Kimia Sains dan AplikasiJournal of Scientific and Applied Chemistry

Journal homepage: http://ejournalundip.ac.id/index.php/ksa

An Insight of Co-Encapsulation Nigella sativa and Cosmos caudatusKunth Extracts as Anti-Inflammatory Agent Through In Silico Study

H)a,b,*Nadiyah Zuhroha, Zubaidah Ningsiha, Anna SafitriCheck forupdates

aChemistry Department, Brawijaya University, Malang, Indonesiab Research Centre of for Smart Molecules of Natural Resources (SMONEGENES), Brawijaya University, Malang, Indonesia

* corresponding author: a.safitri@ub.ac.id

https://doi.0rg/10.14710/ jksa.24.5.152-160

Ar t i c l e In fo Abs t r ac t

Article history:

Received: 24th April 2021Revised: 26th June 2021Accepted: 28th June 2021Online: 31st July 2021

This study analyzes anti-inflammatory activity from extracts of Nigella sativa andCosmos caudatus Kunth co-encapsulated through in silico molecular docking. TheLC-MS results revealed that extracts of N. sativa mostly contained thymoquinoneand alpha-hederin , whereas quercetin and kaempferol were the majorcompounds in C. caudatus K. Nevertheless, the bioactive compounds are usuallysusceptible to degradation by exposure to light, heat, oxygen, which may limit itsbiological activity. Therefore, encapsulation is one of the promising techniques toprotect bioactive compounds. Ligands were encapsulated with chitosan andsodium tripolyphosphate as wall materials. Cyclooxygenase-1 (COX-i) andcyclooxygenase-2 (COX-2) as the target enzymes were docked with acombination of these active compounds (non-encapsulated and encapsulated) ,using the HEX 8.0 program, and visualized using the Discovery studio visualizersoftware vi6.1.0.15350. Interestingly, docking results of the combination ofencapsulated ligands showed no interactions to COX-i but interacted with COX-2. Therefore, co-encapsulation of extracts combinations has been suggested toact as anti-inflammatory agents targeted specifically to the COX-2 enzyme. Thetotal energy of the encapsulated of combination of extract compounds to COX-2were -1425.88 (mol/cal) for thymoquinone + quercetin; -1435.87 (mol/cal) forthymoquinone + kaempferol; 1175.97 (mol/cal) for quercetin + alpha hederin; -957.74 (mol/cal) for kaempferol + alpha hederin; and -283.3 (mol/cal) fordiclofenac sodium, as a control NSAID drug. These suggest that encapsulatedactive compounds in N. sativa and C. caudatus K. have potency as a drug candidatefor the selective NSAIDs category, which can be subjected to further in vitro and invivo studies.

Keywords:

COX-2; co-encapsulation;Cosmos caudatus Kunth; In silico;Nigella sativa

extensive, especially in tropical regions, i.e., Indonesia [5,6]. The bioactivity of the active compounds in C. caudatusKhas great benefits, including antioxidant, antibacterial,antiedema, antifungal, anti-inflammatory, anti-tumor,and antiviral [7, 8].

The stability of natural bioactive compounds, i.e.,preservation of their functional properties, could beimproved using encapsulation techniques, such as spraydrying, spray cooling, coacervation, extrusion, and

1. IntroductionBlack cumin ( N. sativa ) is an herbal plant found in the

Mediterranean and Southeast Asian countries, includingIndonesia. The active compounds in black cumin haveanticancer, antioxidant, and anti-inflammatory activity[1, 2, 3, 4]. Apart from black cumin, another medicinalplant with various benefits isC. caudatus K., locally knownas kenikir. C. caudatus K. is a plant originating from theplains of America, and currently, its distribution is

Jurnal Kimia Sains danAplikasi 24 (5) (2021):152-160 153

polymerization [9, 10]. Encapsulation is the process ofcoating the core material in solid, liquid, or gas particlesusing a coating material. This process aims to protect thecore material in bioactive compounds from variousenvironmental influences such as light, oxygen, water ,and temperature [9,10]. The encapsulation technique canalso increase the absorption of the drug when enteringthe body; hence, drugs can function optimally [10]. Thecoatings on encapsulation can be conducted using theionic gelation technique, prepared using crosslinkerpolymer [11]. Encapsulation can be carried out by chitosanand sodium tripolyphosphate ( Na-TPP) , which act ascrosslinker and coating agents that will protect the activecompounds [11].

Drug activity in the human body plays an essentialrole in preventing and treating diseases. One of them isthe anti-inflammatory activity of a drug which has a vitalrole in the early prevention of disease. Chronicinflammation is closely related to various diseases, suchas arthritis, atherosclerosis, and cancer [12].Inflammation is the sign of pathological or abnormalityin tissue to alert as a trouble signal to the system. Theinflammatory treatment strategy is to involve non-steroid drugs that act on the arachidonic acid pathway byinhibiting both cyclooxygenase-i (COX-i) andcyclooxygenase-2 (COX-2) enzymes [13, 14]. COX-i ispresent in many tissues and produces prostaglandins thatpredominantly regulate normal cellular activities [15].COX-2 activity is usually undetectable in most tissues butcan be rapidly induced by proinflammatory cytokines orgrowth factors. Inhibition of COX-2 prevents high levelsof local production of prostanoids, which results inreduced pain , edema, inflammation, and fever [16]. Theselective NSAIDs inhibit COX-2 favorably, hence,reducing the unwanted effects of nonselective NSAIDs onthe upper GI tract mucosa [17].

To the best of our knowledge, there are no studies ofco-encapsulation for the combination of extracts from N.sativa and C. caudatus K. This approach allowsmanagement of the biologically active compounds of bothN. sativa and C. caudatus K., in a single formulation, withmany applications. One of those is as an anti-inflammatory candidate.

As an initial study, in silico analysis, drug candidatesmust be applied to determine their anti-inflammatoryactivity [18,19]. In silico molecular docking demonstratesan interaction between two or more molecules in the moststable state and can be used as an initial examination indetermining drug candidates based on the interaction ofthe proposed drug (ligands) with the drug target [20].This study aims to determine the effect of thecombination of active compounds on N. sativa and C.caudatus K. co-encapsulated with chitosan and Na-TPP ininhibiting COX-i and COX-2 enzymes by in silicomolecular docking to determine their anti-inflammatoryactivity. Molecular docking was carried out on the mainbioactive compounds in the extracts of N. sativa and C.

caudatus K. The presence of the compounds in N. sativaand C. caudatus K. is analyzed with liquidchromatography-mass spectroscopy (LC-MS).

2. Methodology2.1. Extraction and LC-MS Analysis

The N. sativa and C. caudatus K. dried plants wereobtained from Materia Medica, Batu , East Java, Indonesia.The species of the plants was confirmed by the letter ofspecies determination, signed by a taxonomist from theherbarium unit of Materia Medica. The dried plants werecrushed into powder. Ethyl alcohol pro analysis (p.a) waspurchased from Sigma-Aldrich. The extracts of N. sativaand C. caudatus K. were prepared by the macerationmethod. A100 gram of dried plant powder was immersedin 400 mL ethyl alcohol for 3x 24 h.The resultant extractswere filtered using filter paper. The obtained filtrate wasthen concentrated with a rotary evaporator vacuum at50°C. Extracts were stored at 4°C for further analysis.

The extracts from N. sativa and C. caudatus K werecharacterized using LC-MS. Liquid chromatographicseparation was conducted in the analytical andinstrumental chemistry laboratory of the ChemicalEngineering Department, Politeknik Negeri Malang(POLINEMA). Columns used were a Hypersil Gold (50 mmx 2.1 mm x 1.9 pm), UHPLC brand ACCELLA type 1250made by Thermo Scientific, consisting of a vacuumdegasser and a quartener pump, a thermostaticautosampler controlled by a PC through the X-Calibur 2.1program.Solvents used were solvent A = 0.1% formic acidin water and solvent B = 0.1% formic acid in acetonitrile.A mobile phase gradient was applied , with a flowrate of300 pl/min, with the following settings: 0.0-0.6 min 15%B; 0.6-5.0 min 65% B; 3.0-3.5 min 95% B; 3.5-4.0 min95% B; 4.0-4.5 mini5% B and 4.5-6.0 min 15% B. Theinjection volume was 2 pi. The column was controlled at30°C, and the autosampler compartment was set for i6°C.The MS/MSTriple QXquadrupole) mass spectrometer wasTSQ.QUANTUM ACCESS MAX from Thermo Finnigan withESI (electrospray ionization) ionization method. Thesource was controlled by TSQTune software, operated inpositive ion mode. The ESI ionization conditions were asfollows: 3 kV spray voltage; evaporation temperature250°C; capillary temperature 300°C; nitrogen as sheathgas pressure at 40 psi, and aux gas pressure 10 psi withargon gas.2.2. Ligands Encapsulation with Chitosan and Na-TPP

The preparation of co-encapsulation of N. sativa andC. caudatus K. extracts wascarried out by weighing 500 mgof extracts mixture in a 1:1 ratio. The extracts were thendissolved in 17.5 mL of ethyl alcohol, then a 50 mL of 1%chitosan solution (v/v) in glacial acetic acid solution wasadded. An aliquot of Na-TPP solution (200 mL, 0.5%) wasadded gradually while stirring at a steady speed for onehour. The microencapsulated colloid of N. sativa and C.

Jurnal Kimia Sains danAplikasi 24 (5) (2021):152-160 154

caudatus K. extracts was freeze-dried until the solutionbecame microcapsules powder.

2.3. Ligands and Receptor Preparation

The structure of ligands was obtained from thePubChem NCBI database. The ligands used werecompounds from N. sativa and C. caudatus K extracts:thymoquinone (CID: 10281), quercetin (CID: 5280343),kaempferol (CID: 5280863), alpha-hederin (73296),chitosan (CID: 71853), sodium tripolyphosphate (CID:24455), and diclofenac sodium (CID: 5018304). Thethree-dimensional structures of ligands were prepared byminimizing the binding energy using PyRx software andchange the SDF format to PDB format. Protein Data Bank(PDB) from https:/ /www.rscb.org was used to acquire adatabase to get the three-dimensional structure of COX-1 (PDB ID: 6Y3C) and COX-2 (PDB ID: 5F19). Water andother ligands bound to receptor target were removedusing Discovery studio visualizer VI6.1.0.15350 software[21].

2.4. Molecular Docking

COX-i and COX-2 enzymes were docked to ligandcombinations from N. sativa and C. caudatus K. extractswithout encapsulation (ai-di). For the encapsulatedligands, ligands combinations were docked to chitosanfirst. Thus, a ligand combination-chitosan was formed.After that, the ligan combination-chitosan complexeswere docked with Na-TPP, resulting in ligandcombinations-chitosan-Na-TPP complexes. Thismicrocapsule complex was then docked to the targetenzymes, COX-i or COX-2. The encapsulated ligands withchitosan and Na-TPP compounds were designated as a2,C2, and d2; and diclofenac sodium as a reference ligandwas also docked to COX-i and COX-2 enzyme anddesignated as e. Molecular docking was established usingHex 8.0.0 software. Analyses of the molecularinteractions of all ligands and receptors were conductedby Discovery studio visualizer vi6.1.0.15350 software [21].The amino acid residues, hydrogen bonds, van der Waalsforces, and energy binding in the interactions betweenligands and receptors were analyzed. The two-dimensional ligands and receptor interactions werevisualized by the LigPlot+ program [22].

3. Results and DiscussionExtracts of N. sativa seed and C. caudatus K. leaves

were characterized by the LC-MS analysis (Figures 1-2).The major compounds in N. sativa extract werethymoquinone and alpha-hederin. C. caudatus K. extractswere mostly contained quercetin and kaempferol as theiractive compounds (Figure 2). The retention time (RT) forthe active compound shows peak of thymoquinone(C10H12O2) at RT = 0.76 min, m/z 164.50-165.50, alpha-hederin (C4IH660I2) had RT = 1.37 min, m/z 751.50-752.50(Figure1).The peak of quercetin (CI5HI007) appeared at RT= 2.70 min, m/z 300.50-301.50; kaempferol (C^HKAS)peak at RT = 2.79 min , m/z 285.50-286.50 (Figure 2).

These mass spectra agreed with the previously publishedliterature reporting of mass spectrometry ofthymoquinone, alpha-hederin [23], quercetin , andkaempferol [24]. The prominent peaks in the activecompounds of each extract resulted from the LC-MSanalysis became the standard for selecting compounds(ligands) in the subsequent in silico study.

The ligands-receptor interaction presented by thebinding sites on amino acid residues and the types offormation of chemical bonds is shown in Figures 3 and 4.Five hydrogen bonds, two electrostatic bonds, and sevenhydrophobic bonds, with the amino acid residuesinvolving Gln44, Gly47i, Lys468, His43, Arg83, Leui23,Tyr64, and Vain9 were shown in the interactions of non-encapsulated thymoquinone + quercetin to COX-i, withthe binding energy of -303.19 cal/mol (Figure 3ai). In theinteractions of non-encapsulated thymoquinone +

kaempferol to COX-i, there were two hydrogen bonds andeight hydrophobic bonds with amino acid residuesArgi20, Valii9, Pro84, Leuii5, and Valn6, and had -299.29 cal/mol binding energy (Figure 3bi). The aminoacid residue Argi20 is located close to the active site of theCOX-i enzyme and is one of the amino acid residues thataffect COX-i inhibition [25]. In Figure 301, the bindingenergy of -406.53 cal/mol resulted in a complex of non-encapsulated quercetin + alpha hederin, with fourhydrogen bonds and ten hydrophobic bonds formed fromLeuii5, Leui23, Arg83, Valii9, Arg79, and Tyr64 (Figure3ci). Four hydrogen bonds and one Pi-sigma bond in anon-encapsulated kaempferol + alpha-hederininteracted with Gly552, and the binding energy was -177.40 cal/mol (Figure 3di). Interestingly, whenencapsulated ligands and diclofenac sodium were dockedto COX-i, as shown in Figures 3a2, 3b2, 3C2, 3d2, and 3e,there were no bonds formed between those ligands andreceptor target, which were also confirmed by the two-dimensional conformation. The value of the resultedbinding energy from the molecular dockings may occurfrom the presence of bonds within ligands and thesolvent's influence during the docking process [26].a 0.76100

g80u

51 60-5f0

2 x 2.43 2.89 3.32 349 587® 40- 0 24

& 2(Y3.88 5.782.02 4.131.64 4 55 5.55

b 0.48100- 0.67gS 80S? 60

0.71 1.35 1.375re

1.44l& 20

0.79S 1.491-58 2.03 2.64 3.38 3.84 4.57 4.79 5.36 5.78

Time (min)

Figure l. The LC-MS results from N.sativa seed extract:(a) thymoquinone; (b) alpha-hederin.

Jurnal Kimia Sains danAplikasi 24 (5) (2021):152-160 155

interactions of non-encapsulated thymoquinone +

quercetin to COX-2 (Figure 4ai). In the interactions ofnon-encapsulated thymoquinone + kaempferol to COX-2, three hydrogen bonds and ten hydrophobic bonds withthe amino acid residues involved His388, Gln203, His386 ,Leu294, Val29i, Val447, His207, and His2i4 (Figure 4bi).Figure 4ci (quercetin + alpha-hederin) showed sevenhydrogen bonds and six hydrophobic bonds with aminoacid residues involving His35i > Ser58i, Ser579 Asp347 ,Gly354, GIU346, Lys358, Pro5i4, His95, and His356(Figure 4ci). Four hydrogen bonds, two electrostaticbonds, and six hydrophobic bonds with the amino acidresidues involved Gln370, Hisi22, Seri26, Arg6i, Proi27,Lysi37, and Trpi39, the interaction of non-encapsulatedkaempferol + alpha-hederin interacted with COX-2(Figure 4di). The binding energies of encapsulatedthymoquinone + quercetin, thymoquinone + kaempferol,quercetin + alpha-hederin, and kaempferol + alpha-hederin to COX-2 enzyme were -331.10 cal/mol, -283.03cal/mol, -418.20 cal/mol, and -402.92 cal/mol,respectively.

2.70a g 100.8 80<0|60

40fI 200

0.53r 0.73088 141

2.11> 2.88 3 22 3.48 4.16 4.70 4.72 5.73ct

b 2.79* 100 J

s 80re

1 6(H

I *3 0.720.68is 0.97 4.393.021.13£ 20 ;0.39 1.50 2.09 3.20 4 05 4 52 4.86 5.75-fcTime (min)

Figure 2. The LC-MS results from Cosmos caudatus Kleaves extracts: (a) quercetin; (b) kaempferol.

The interaction of both non- and encapsulatedligands to COX-2 enzyme is shown in Figure 4, and thetype of chemical bonds formed is listed in Table 1. Therewere four hydrogen bonds and nine hydrophobic bonds,with amino acid residues involving His207, Gln454,His2i4, Val 291, Val447, His386, and His388 in the

Energy( mol/cal)

EnergyNon-Encapsulation-COX-1 Eneapsulation-COX-1

(mol/cal)

(a2)(al )

-484.05'".' i ' -303.19

A

-299.29 -449.29

-406.53

..

(d l ) (d2)

-177.40 -702.64

Figure 3. Molecular interaction between COX-i non-encapsulated (1) and encapsulated (2) ligand combination from:(a) thymoquinone + quercetin; (b) thymoquinone + kaempferol; (c) quercetin + alpha-hederin; (d) kaempferol +

alpha-hederin; and (e) diclofenac sodium. Encapsulated ligands are when ligands are docked to chitosan and Na-TPP,then docked to enzyme target.

Jurnal Kimia Sains danAplikasi 24 (5) (2021):152-160 156

Energy(mol/cal)

Energy(mol/cal)

Non-Encapsulation-COX-2 Eneapsnlation-COX-2

-1425.88

( bl ) (b2)

-283.03 -1435.87Xv I <1141 NA)

<3W00(.\ )

IIKIIWA"; "£(Cl)

-418.20 -1175.97-f :

-402.92 -957.74

(e)

n* . i

$ IV l l. KMtu l <2m -283.3< >*» U *)

Figure 4. Molecular interaction between COX-2 non-encapsulated (1) and encapsulated (2) ligand combination from:(a) thymoquinone + quercetin; (b) thymoquinone + kaempferol; (c) quercetin + alpha-hederin; (d) kaempferol +

alpha-hederin; and (e) diclofenac sodium. Encapsulated ligands are when ligands are docked to chitosan and Na-TPP,then docked to enzyme target.

In the encapsulated ligands against COX-2, as shownin Figures 4a2-4d2, lower binding energies (morenegative) than those in the interaction between non-encapsulated ligands and COX-2. This means thatencapsulated ligands interacted stronger with COX-2than non-encapsulated ligands to COX-2.

There were seven hydrogen bonds, two electrostaticbonds, and other bonds (Pi-lone pair type) , with aminoacid residues involving Asp258, Asp399, GIU416, Leu4i5,and His282 in the encapsulated thymoquinone +

quercetin docked to COX-2 (-1425.88 cal/mol). Therewere two hydrogen bonds, four electrostatic bonds, andtwo hydrophobic bonds with amino acid residuesinvolving GIU516, GIU424, His4i7, Tyr402, GIU416, andHis398 in the interactions of encapsulated thymoquinone+ kaempferol to COX-2, with -1435.87 cal/mol bindingenergy (Figure 4bi). When encapsulated quercetin + alphahederin bound COX-2, the binding energy was -1175.97cal/mol, resulted in seven hydrogen bonds, fiveelectrostatic bonds, and one hydrophobic bond with the

amino acid residues involved Asp399, Glu4i6, GIU424,Gln400, Leu4i5, Asp258, Thr420, and His4i7 (Figure4c2). In Figure 4d2, encapsulated kaempferol + alphahederin had formed three hydrogen bonds, twoelectrostatic bonds, one hydrophobic bond, and twometal-acceptor type bonds to the amino acid residuesinvolving Glu4i6, His398, Asp399, Val295, Phe200, andIle4i3, and had -957.74 cal/mol binding energy.

Interaction between a control ligand, diclofenacsodium, and COX-2 resulted in 6 hydrophobic bonds andtwo metal-acceptor bonds in the amino acid residuesSeri2i, Ilei24, Arg469, Cys4i, Arg44, Leui52, Arg469,and Leui52. Intriguingly, the binding energy of diclofenacsodium to COX-2 was higher than those of allcombinations of encapsulated ligands at -283.3 cal/mol,suggesting that encapsulated ligands have moreinteractions to COX-2 than diclofenac sodium.Nevertheless, sodium diclofenac is one of thenonselective NSAIDs with the utmost COX-2 selectivity[18].

Jurnal Kimia Sains danAplikasi 24 (5) (2021):152-160 157

Table 1. Ligand interaction of COX-2 with non-encapsulated (1) and encapsulated (2) ligand combination of: (a)thymoquinone + quercetin; (b) thymoquinone + kaempferol; (c) quercetin-alpha hederin; (d) kaempferol + alpha-

hederin; and (e) diclofenac sodium.Label Interaction Chemistry bond Label Interaction Chemistry bond(ai) A:HIS207:HDl - :LIG:0

A:GLN454:HE22 - :LIGl:0A:HIS207:CEl - :LIGl:0

A:HIS2l4:CEl - :LIGl:0A:VAL29l:CGl - :LIGlA:VAL291:CG2 - :LIGl:LIGl:C- a:VAL447A:HIS207 - :LIGl:C

A:HIS2l4- :LIGl:CA:HIS386 - :LIGl:C

A:HIS388 - :LIGl:C:LIGl - A:VAL447

Hydrogen bondHydrogen bondHydrogen bondHydrogen bondHydrophobicHydrophobicHydrophobicHydrophobicHydrophobicHydrophobicHydrophobicHydrophobic

(a2) :UNKl:P6 - A:ASP258:OD2

:UNKl:PlO - A:ASP258:OD2

:UNKl:H - A:ASP399:OD2

:UNKl:H - A:ASP258:OD2

:UNKl:H - A:GLU4l6:OE2

:UNKl:H - A:LEU4l5:0:UNKl:09l - A:HIS282

Electrostatic

ElectrostaticHydrogen bondHydrogen bondHydrogen bondHydrogen bond

Other bond ( Pi-lone pair type)

(bl) :LIG:H - A:HIS388:NE2:BA:GLN203:HE2l - :LIGl:0A:HIS386:CD2- :LIGl:0A;LEU294:CD2 - :LIGl

:LIGl:C- A:VAL29l:LIGl - A:VAL447:LIGl - a:VAL29l

A:HIS207 - :LIGl:CA:HIS214 - :LIGl:CA:HIS386 - :LIGl:CA:HIS388 - :LIGl:C

Hydrogen bondHydrogen bondHydrogen bondHydrophobicHydrophobicHydrophobicHydrophobicHydrophobicHydrophobicHydrophobicHydrophobic

(b2) :UNKl:H - A:HIS417:0:UNKl:H - A:TYR402:OH

:UNKl:NAl5 - A:GLU4l6:OE2

:UNKl:NAl7 - A:GLU424:OE2

A:GLU4l6:OEl - :UNKl

A:GLU4l6:OE2 - :UNKl

:UNKl:H - A:HIS417A:HIS398 - UNKl:Cl

Hydrogen bondHydrogen bond

ElectrostaticElectrostaticElectrostaticElectrostaticHydrophobicHydrophobic

(cl) A:HIS35l:HN - :LIGl:050A:HIS35l:HDl - :LIGl:0A:SER58l:HG - :LIGl:0:LIGl:H - A:SER579:OG:LIGl:H - A:ASP347:0

A:GLY354:CA - :LIGl:0:LIGH57 " A:GLU346:0

:LIGlCl - A:LYS358:LIGl:C4l - A:PR0514A:HIS95 - :LIGl:C42

A:HIS356 - :LIGlA:HIS356 - :LIGl:C48

Hydrogen bondHydrogen bondHydrogen bondHydrogen bondHydrogen bondHydrogen bondHydrogen bondHydrophobicHydrophobicHydrophobicHydrophobicHydrophobic

(C2) A:GLN400:HE22 - :UNKl:0

:UNKl:H - A:LEU4l5:0:UNKl:H - A:ASP399:ODl

:UNKl:H - A:ASP258:OD2

:UNKl:N95 - A:HIS4l7:UNKl:NAl5 - A:ASP399:OD2

:UNKl:NAl5 - A:GLU4l6:OE2

:UNKl:NAl6 - A:ASP399:ODl

:UNKl:NAl7 - A:GLU424:OElA:HIS417 - :UNKl:C4l

Hydrogen bondHydrogen bondHydrogen bondHydrogen bond

ElectrostaticElectrostaticElectrostaticElectrostaticElectrostaticHydrophobic

(dl) :LIGl:H58 - A:GLN370:0:LIGl:H59 - A:GLN370:0

:LIGl:H63- A:HISl22:0:LIGl:H77 - A:SERl26:0

A:ARG6l:NH2 - :LIGlA:PROl27 - :LIGl

:LIGlC4l - A:LYS137:LIGl:C47 " A:PROl27A:TRPl39 - :LIGl:C42

Hydrogen bondHydrogen bondHydrogen bondHydrogen bond

ElectrostaticHydrophobicHydrophobicHydrophobicHydrophobic

(d2) A:HIS398:HDl - :UNKl:024:UNKl:H - A:GLU4l6:OEl

:UNKl:H - A:ASP399:OD2

:UNKl:NAl5 - A:GLU4l6:OE2

:UNKl:NAl6 - A:GLU4l6:OEl:UNKl:Cl - A:ILE413

Hydrogen bondHydrogen bondHydrogen bond

ElectrostaticElectrostaticHydrophobic

:UNKi:Nai7- A:VAL295:0 Other bond (Metal- acceptor type)

:UNKi:Nai8- A:PHE200:0 Other bond (Metal- acceptor type)

(e) :UNK:CLl8- A:ARG469:UNK:CL19 - A:CYS4l

:UNKl - A:ARG44:UNKl - A:LEU152:UNKl - A:ARG469

HydrophobicHydrophobicHydrophobicHydrophobichydrophobic

:UNKi:NA20 - A:SERi2i:0 Other bond (metal-acceptor type)

:UNKi:NA20 - A:ILEi24:0 Other bonds (metal-acceptor type)

between

ligands and complex with solvents, with the binding energy calculation

derived from Δ𝐺 = 𝐺 complex - [𝐺 receptor + 𝐺 ligand] .

Nevertheless

β

Jurnal Kimia Sains danAplikasi 24 (5) (2021):152-160 158

Molecular docking is a widely applied, rapid, andcost-effective computational tool for predicting in silicothe binding models and affinities of molecularidentification events [26, 27, 28]. Receptor-liganddocking represents a predominantly important techniquebecause of its importance in the current drug discoverydevelopment [29]. An in-depth understanding of thenature of molecular recognition is also of greatsignificance in facilitating the discovery, design, anddevelopment of novel drugs. In the current study, throughin silico molecular docking, encapsulation of ligandcombination from N. sativa and C. caudatus K. extracts isproposed to have selective anti-inflammatory activitytoward COX-2 enzyme.

There are some possible explanations for theseresults. First, the active sites in COX-2 are larger thanCOX-i. Hence the extension in ligand sizes, as in the caseof encapsulated ligands, can increase the selectivitytoward COX-2 enzyme [30]. Another thing is that sincethe encapsulated ligands are bulky and more complex, asa result , those ligands have more hydrophobiccharacteristics. Ligands with hydrophobic interactionscontribute significantly to the binding affinity of thedocked complex and are entropy-driven interactions[28].

AcknowledgmentThe Indonesian Endowment Fund for Education/

Lembaga Pengelola Dana Pendidikan ( LPDP) from theIndonesian Ministry of Finance has provided financialsupport for this study.

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In this current work, encapsulated ligands had lowerbinding energy than non-encapsulated ligands wheninteracting with the COX-2 enzyme, indicating a strongerinteraction with the enzyme [20]. The binding affinityvalue comes from the interaction between the ligandsandthe target compounds and can occur from interactionwithin ligands and the solvent 's influence during thedocking process [26]. The total energy produced bydockings ligands-receptor can be influenced by some ofthe existing energies such as energy in the Van Der Waalsforces, electrostatic bonds, or energy that occurs

[31], the molecular docking results should be

investigated further through in vitro and in vivoapproaches.

4. ConclusionThymoquinone and alpha-hederin are suggested to

be the primary compounds in N. sativa extract. Moreover,C. caudatus K. extract compounds were mainly quercetinand kaempferol. The in silico molecular docking on thecombination of compounds in N. sativa and C. caudatus K.showed that encapsulated ligands possess specific anti-inflammatory through inhibition of COX-2 enzyme. Inaddition, encapsulated ligands had a stronger ligand bondwith the receptor target, as indicated by the smallerenergy produced in the encapsulation results. Thissuggests that encapsulated active compounds in N. sativaand C. caudatus K can be proposed for selective anti-inflammatory drugs.

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