|Year : 2021 | Volume
| Issue : 2 | Page : 169-174
Potential anti-alopecia constituents from Theobroma cacao: An in silico study
Dikdik Kurnia1, Faruk Jayanto Kelutur2, Resmi Mustarichie2
1 Department of Chemistry, Faculty of Mathematics and Natural Sciences, Padjadjaran University, Sumedang, West Java, Indonesia
2 Department of Pharmaceutical Analysis and Medicinal Chemistry, Faculty of Pharmacy, Universitas Padjadjaran, Sumedang, West Java, Indonesia
|Date of Submission||16-Dec-2020|
|Date of Decision||17-Feb-2021|
|Date of Acceptance||03-Mar-2021|
|Date of Web Publication||27-Apr-2021|
Dr. Dikdik Kurnia
Department of Chemistry, Faculty of Mathematics and Natural Sciences, Universitas Padjadjaran, Jl. Raya Bandung Sumedang KM. 21, Jatinangor, West Java 453
Source of Support: None, Conflict of Interest: None
Tinea capitis is local alopecia caused by a dermatophyte infection of the scalp. Trichophyton rubrum produces the squalene epoxidase enzyme, which has a crucial role in prolonged dermatophyte infection, as well as in synthesizing fatty acids in this dermatophyte group. This study analyzes Trichophyton cacao compounds as anti-alopecia by inhibiting the squalene epoxidase enzyme formation, in silico. The structure of T. cacao compounds was prepared using the MolView Web application. The compound docked to squalene epoxidase using AutoDock Vina in PyRx 0.8, followed by PyMOL for visualization, and the Proteins Plus program to analyze the complexity. The binding affinity value of catechin, epicatechin (−8.0 kcal/mol), and anthocyanin (−7.8 kcal/mol) compounds was higher than the positive control (terbinafine, −6.7 kcal/mol). Pre-ADMET demonstrated that catechin and epicatechin had moderate Human Intestinal Absorption (66.71%), but anthocyanin was very good (100%). Caco-2 parameters for catechin and epicatechin were relatively low (<4 nm s− 1), while anthocyanin, theobromine, and terbinafine were within 4–70 nm s− 1. Plasma protein binding shows catechin, epicatechin, and anthocyanin diffuse through the plasma membrane and interact with plasma proteins. The toxicity results for all compounds are mutagenic, and only terbinafine is carcinogenic. Based on the Lipinski's “Rule of Five,” compounds from T. Cacao can be given orally. Catechin and epicatechin compounds have the potential to act as anti-alopecia. These two compounds can diffuse and interact with plasma proteins so they are directly on the target when given orally.
Keywords: Alopecia, squalene epoxidase, Trichophyton cacao compounds, Trichophyton rubrum, tinea capitis
|How to cite this article:|
Kurnia D, Kelutur FJ, Mustarichie R. Potential anti-alopecia constituents from Theobroma cacao: An in silico study. J Adv Pharm Technol Res 2021;12:169-74
|How to cite this URL:|
Kurnia D, Kelutur FJ, Mustarichie R. Potential anti-alopecia constituents from Theobroma cacao: An in silico study. J Adv Pharm Technol Res [serial online] 2021 [cited 2021 Jun 25];12:169-74. Available from: https://www.japtr.org/text.asp?2021/12/2/169/314681
| Introduction|| |
Hair has an important role in protecting the scalp from the surrounding environment, and influences one's esthetic appearance and self-confidence. Hence, hair loss that causes baldness has been shown to lead to a frustrating effect. Hair can show a group of people's identities, whether based on ethnicity or culture. Thus, hair loss must be diagnosed early to treat its causes more effectively.
Alopecia is a condition in which hair loss in the scalp area and other body parts is excessively abnormal. In general, a large volume occurs on the scalp. Types of alopecia consist of scarring alopecia (associated with fibrosis, inflammation, and loss of hair follicles) and nonscarring alopecia (associated with hair follicles present, although may miss some hair strands). External factors that cause nonscarring alopecia are tinea capitis, alopecia areata, traction alopecia, and trichotillomania.,,
Tinea capitis is local alopecia caused by a dermatophyte infection around the scalp and hair shaft, accompanied by squama. In general, this type of alopecia occurs in children. It is alopecia that is caused by Trichophyton rubrum, a dermatophyte that infects the skin's superficial layer by eating any keratinized areas. T. rubrum produces the squalene epoxidase enzyme, which plays an important role in synthesizing fatty acids in its dermatophytes and sterol biosynthesis (ergosterol), so that the enzyme becomes the primary target in the inhibition of dermatophytosis.
The drug that has been commercially used to treat dermatophytosis is terbinafine, N, 6,6-trimethyl-N-(naphthalene-1-ylmethyl) hept2-en-4-yn-1-amine, which works by inhibiting activity of the squalene epoxidase enzyme so that ergosterol synthesis is disrupted and fungal cells die. However, the use of these drugs is inefficient in the long term. There is a risk of increasing serotonergic effects, primarily due to reduced analgesia associated with l-opioid receptors. Therefore, the design and development of a new drug are needed.
The use of natural materials as drug ingredients is essential. Trichophyton cacao contains polyphenol compounds (catechin, epicatechin, anthocyanin, phenolic acid, tannin, proanthocyanidin flavonoid, and others). It has been reported that the most components in this natural material, namely, catechin and epicatechin, have anti-carcinogenic, anti-inflammatory, anti-microbial, and antioxidant activities. Therefore, further research is needed to investigate promising anti-alopecia agents. This study was designed to predict anti-alopecia compounds' molecular action on T. cacao by inhibiting dermatophytes that can form tinea capitis. The in silico method was employed to investigate the ability to predict potential target proteins that bind these compounds.
| Materials and Methods|| |
The test ligands from T. cacao compounds (catechin, anthocyanin, theobromine, and epicatechin) were prepared using the MolView program (http://molview.org/) and positive control; terbinafine was obtained from PubChem (https://pubchem.ncbi.nlm.nih.gov/) with CID, 1549008. The squalene epoxidase target enzyme used in this study was obtained from UniProt (http://www.uniprot.org/) with the gene name Q4JEX9.
Preparation of test ligands and target enzyme
The test ligands for the T. cacao compounds (catechin, anthocyanin, theobromine, and epicatechin) were prepared using the MolView program and then stored in the MOL file format. The OPEN BABEL 2.4.1 program was used to change the positive control (terbinafine) to the PDB file format. The target enzyme (squalene epoxidase) was prepared using the SWISS-MODEL (https://swissmodel. expasy. org/), and converted to the PDB file format.
Molecular docking of Theobroma cacao compounds against squalene epoxidase
The docking process was carried out using AutoDock Vina software, which is connected to the PyRx 0.8 tools., The target enzyme, squalene epoxidase, was selected for the macromolecule. The test ligands from T. cacao compounds and positive control were docked at the active sites of squalene epoxidase using PyRx. The result obtained is binding affinity (kcal/mol) to compare each protein-ligand complex value. The conformation was chosen based on the lowest binding affinity, which has a value of Root Mean Square Deviation (RMSD) ≤1.0 Å.,
Visualization and analysis of molecular docking results
PyMOL was used to visualize poses best when docking and protein-ligand molecular interactions. Meanwhile, Protein Plus (https://proteins.plus/) used to analyze the amino acid residues formed in the form of two dimensional (2D) and 3D structure images. The similarity can be seen of ligand poses to one another on the target protein's active sites when bound.
Pre-ADMET and lipinski rule of five
The pre-ADMET program was accessed through http://preadmet.bmdrc.kr/adme, with the chemical structures of the compounds, described or uploaded in Mol file format (*.mol). The program automatically calculates the value of the selected parameter, including human intestinal absorption (HIA), cell permeability of human colon adenocarcinoma (Caco-2), plasma protein binding (PPB), mutagenic, and carcinogenic. Lipinski's “Rule of Five” was accessed through scfbio-iitd.res.in/software/drugdesign/lipinski. jsp with the Protein Data Bank file format (*.pdb). The parameters observed were molecular weight, log P, hydrogen bond donor and acceptor, and molar refractivity.
| Results|| |
Results of molecular docking of Theobroma cacao compounds against squalene epoxidase enzyme
[Table 1] shows the binding affinity and interaction between the ligands and the squalene epoxidase.
|Table 1: Prediction of anti-alopecia activity from Theobroma cacao compounds|
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[Table 2] shows that the compound structure from T. cacao is not much different from the positive control (terbinafine).
|Table 2: Structure and a molecular formula of the Theobroma cacao compounds and positive control|
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Visualization and analysis of the molecular docking results between the ligand and squalene epoxidase in [Figure 1] and [Figure 2] demonstrated the active sites' binding.
|Figure 2: Visualization of catechin (a), anthocyanin (b), epicatechin (c), theobromine (d), and terbinafine (e) bound to the active sites of squalene epoxidase|
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Predictions of absorption, distribution, metabolism, excretion, and toxicity are shown in [Table 3].
|Table 3: Preabsorption, distribution, metabolism, excretion, and toxicity prediction results|
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Lipinski rule of five
The predictive route of oral drug administration is shown in [Table 4].
|Table 4: Prediction results of compounds based on the Lipinski's rule of five|
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| Discussion|| |
Prediction of bioavailability and anti-alopecia activity from Theobroma cacao compounds through molecular interaction with squalene epoxidase
Binding affinity shows the amount of energy released by a compound (ligand) when it interacts or binds to a target (receptor). In general, a high-affinity ligands' binding result was obtained from a greater intermolecular force between the ligands, which shows good activity., The smaller the value, the stronger the energy used to form a bond. Based on the molecular docking results in [Table 1], the compounds with strong bonds are catechin = epicatechin > anthocyanin > theobromine, with binding affinity values of −8.0, −8.0, −7.8, and −6.1 kcal/mol, respectively. Compared to the positive control (terbinafine,-6.7 kcal/mol), the catechin, epicatechin, and anthocyanin compound better bond to the squalene epoxidase enzyme, so that it can be predicted that the three compounds contained in T. Cacao have potent activities as anti-alopecia. In addition, the RMSD value of the three potent compounds is 0, which indicates a stable position when interacting, and is strongly bound to the substrate (squalene epoxidase enzyme). The interaction's stability is influenced by the distance and hydrogen bonds formed between the ligand and protein, so that the smaller the distance, the stronger the bond energy.
In the development of a new drug substance, absorption, distribution, metabolism, excretion, and toxicity must be thoroughly investigated before clinical testing is carried out. Predictable parameters include pharmacokinetic properties and toxicity predictions. Pharmacokinetic properties consist of absorption (HIA and permeability of human colon adenocarcinoma/Caco-2 cells) and distribution (PPB). Meanwhile, toxicity was done through the prediction of mutagenic and carcinogenic properties.
HIA shows the percentage of bioavailability and absorption evaluated by excretion ratio through urine, bile, and feces. Meanwhile, Caco-2 has been widely used in vitro modeling to predict drug transport through human colon adenocarcinoma's intestinal epithelium, which has multiple transport routes. Distribution parameters pay attention to PPB and are closely related to drug disposition ability when giving effect. In general, only drugs in the unbound form can diffuse through the membrane and interact with pharmacological targets, so plasma protein bonds are crucial in drug efficiency. PPB is a fraction of the available drug-free of charge for distribution to various tissues.
The catechin and epicatechin compounds have a moderate HIA pharmacokinetics value (66.71%) during absorption in the intestine, in contrast to the very good anthocyanins (100%). The Caco-2 parameters showed very low catechin and epicatechin (<4 nm s− 1) in vitro cell permeability. Meanwhile, anthocyanins, theobromine, and terbinafine compounds have moderate permeability (4–70 nm s− 1). In terms of PPB, the catechin, epicatechin, and anthocyanin compounds can diffuse through the plasma membrane and interact with plasma proteins. However, the toxicity results demonstrate that all compounds were mutagenic, and only terbinafine was carcinogenic.
Solubility and permeability predictions based on the Lipinski rule of five
The solubility and permeability of a compound play a crucial role in considering drug development to the next stage. This was done to prevent the drug's failure by low absorption or permeation. Based on the rules of Lipinski on the discovery and development of a candidate drug substance used orally, it must satisfy the five conditions known as the “Rule of Five,” including having a molecular weight value of not more than 500 daltons, having high lipophilicity, which is indicated by a log P of not more than 5, a hydrogen bond donor not more than 5, a hydrogen bond acceptor not more than 10, and a molar refractivity value between 40 and 130. Based on these rules [Table 4], all T. Cocoa compounds can be administered orally.
| Conclusions|| |
The results of molecular docking using the in silico method of T. cacao compounds as anti-alopecia in inhibiting the formation of the squalene epoxidase enzyme have concluded to have a binding affinity value, namely, catechin = epicatechin (−8.0 kcal/mol) > anthocyanin (−7.8 kcal/mol), compared to terbinafine (−6.7 kcal/mol), as a drug that has been used commercially. The Pre-ADMET (pharmacokinetics and toxicity) showed that the catechin and epicatechin compounds had moderate HIA (66.71%) when there was absorption in the intestine, but it was perfect anthocyanin (100%). Caco-2 parameters showed low catechin and epicatechin (<4 nm s− 1) in-vitro cell permeability, but anthocyanin, theobromine, and terbinafine had moderate permeability (4–70 nm s− 1). The PPB for catechin, epicatechin, and anthocyanin compounds can diffuse through the plasma membrane and interact with plasma proteins. However, the toxicity results indicate that all compounds were mutagenic, and only terbinafine was carcinogenic. The Lipinski Rule of five indicates that the T. Cacao compounds can be given orally.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
| References|| |
Jain R, Monthakantirat O, Tengamnuay P, De-Eknamkul W. Identification of a new plant extract for androgenic alopecia treatment using a non-radioactive human hair dermal papilla cell-based assay. BMC Complement Altern Med 2016;16:18.
Birngruber CG, Verhoff MA. Handbook of Hair in Health and Disease. The Netherlands: Wageningen Academic Publishers; 2012. p. 1-493.
Sarifakioglu E, Yilmaz AE, Gorpelioglu C, Orun E. Prevalence of scalp disorders and hair loss in children. Cutis 2012;90:225-9.
Arck PC, Handjiski B, Peters EM, Peter AS, Hagen E, Fischer A, et al
. Stress inhibits hair growth in mice by induction of premature catagen development and deleterious perifollicular inflammatory events via neuropeptide substance P-dependent pathways. Am J Pathol 2003;162:803-14.
Shapiro J, Wiseman M, Lui H. Practical management of hair loss. Can Fam Physician 2000;46:1469-77.
Mulinari-Brenner F, Bergfeld WF. Hair loss: Diagnosis and management. Cleve Clin J Med 2003;70:705-12.
Richardson MD, Warnock DW. Fungal Infection: Diagnosis and Management. 4th
ed. UK: John Wiley & Sons, Ltd; 2011.
Wang W, Sheng C, Che X, Ji H, Miao Z, Yao J, et al
. Design, synthesis, and antifungal activity of novel conformationally restricted triazole derivatives. Arch Pharm (Weinheim) 2009;342:732-9.
Favre B, Ryder NS. Characterization of squalene epoxidase activity from the dermatophyte Trichophyton rubrum
and its inhibition by terbinafine and other antimycotic agents. Antimicrob Agents Chemother 1996;40:443-7.
Nowosielski M, Hoffmann M, Wyrwicz LS, Stepniak P, Plewczynski DM, Lazniewski M, et al.
Detailed mechanism of squalene epoxidase inhibition by terbinafine. J Chem Inf Model 2011;51:455-62.
Petranyi G, Ryder NS, Stütz A. Allylamine derivatives: New class of synthetic antifungal agents inhibiting fungal squalene epoxidase. Science 1984;224:1239-41.
Jain PK, Dass DJ. Evaluating hair growth potential of some traditional herbs. Asian J Pharm Clin Res 2015;8:150-2.
Saarikoski T, Saari TI, Hagelberg NM, Backman JT, Neuvonen PJ, Scheinin M, et al.
Effects of terbinafine and itraconazole on the pharmacokinetics of orally administered tramadol. Eur J Clin Pharmacol 2015;71:321-7.
Kaushik R, Institutions R, Gupta D, Institutions R. Alopecia: Herbal remedies. Int J Pharm Sci Res 2014;2:1631-7.
Arlorio M, Coïsson J, Travaglia F. Antioxidant and Biological Activity of Phenolic Pigments from Theobroma cacao
Hulls Extracted with Supercritical CO2. Food Res Int 2005;38:1009-14.
Hii C, Law C, Suzannah S, Cloke M. Polyphenols in Cocoa
L.). J Food Ag-Ind 2009;2:702-22.
Karumuri S, Singh PK, Shukla P. In silico
analog design for terbinafine against Trichophyton rubrum
: A preliminary study. Indian J Microbiol 2015;55:333-40.
Dallakyan S, Olson AJ. Small-molecule library screening by docking with PyRx. Methods Mol Biol 2015;1263:243-50.
Alamzeb M, Omer M, Ur-Rashid M, Raza M, Ali S, Khan B, et al
. NMR, novel pharmacological and in silico
docking studies of oxyacanthine and tetrandrine: bisbenzylisoquinoline alkaloids isolated from Berberis glaucocarpa
Roots. J Anal Methods Chem 2018;2018:1-7.
Rauf MA, Zubair S, Azhar A. Ligand docking and binding site analysis with PyMOL and AutoDock Vina. Int J Basic Appl Sci 2015;4:168-77.
Mazur M, Olczak J, Olejniczak S, Koralewski R, Czestkowski W, Jedrzejczak A, et al
. Targeting acidic mammalian chitinase is effective in animal model of asthma. J Med Chem 2018;61:695-710.
Kelutur FJ, Mustarichie R, Umar AK. Virtual screening kandungan senyawa kipas laut (Gorgonia mariae) sebagai Antiasma. J Penelit Kim 2020;16:199-210.
Lipinski CA. Lead and drug-like compounds: The rule-of-five revolution. Drug Discov Today Technol 2004;1:337-41.
Du X, Li Y, Xia YL, Ai SM, Liang J, Sang P, et al
. Insights into protein-ligand interactions: Mechanisms, models, and methods. Int J Mol Sci 2016;17:1-34.
Chu W, Prodromou R, Day KN, Schneible JD, Bacon KB, Bowen JD, et al
. Peptides and pseudopeptide ligands: A powerful toolbox for the affinity purification of current and next-generation biotherapeutics. J Chromatogr A 2021;1635:1-68.
Umamaheswari M, Madeswaran A, Asokkumar K. Virtual screening analysis and in vitro
xanthine oxidase inhibitory activity of some commercially available flavonoids. Iran J Pharm Res 2013;12:317-23.
Kapitan OB, Ambarsari L, Falah S. Inhibition docking simulation of zerumbone, gingerglycolipid b, and curzerenone compound zingiber zerumbet from timor island against mura enzyme. J Appl Chem Sci 2016;3:279-88.
O'Hagan S, Kell DB. The apparent permeabilities of Caco-2 cells to marketed drugs: Magnitude, and independence from both biophysical properties and endogenite similarities. Peer J 2015;3:e1405.
Megantara S, Levita J, Iwo MI, Ibrahim S. absorption, distribution and toxicity prediction of andrographolide and its derivatives anti-HIV drugs. Res J Chem Environ 2018;22:82-5.
Nursamsiar, Toding AT, Awaluddin A. Studi in silico
senyawa turunan analog kalkon dan pirimidin sebagai antiinflamasi: Prediksi absorpsi, distribusi, dan toksisitas. Pharmacy 2016;13:92-100.
[Figure 1], [Figure 2]
[Table 1], [Table 2], [Table 3], [Table 4]