Next Article in Journal
Garlic (Allium sativum L.) as an Ally in the Treatment of Inflammatory Bowel Diseases
Next Article in Special Issue
Changes in the Expression Profile of Pyroptosis-Related Genes in Senescent Retinal Pigment Epithelial Cells after Lutein Treatment
Previous Article in Journal
Mapping QTLs for Super-Earliness and Agro-Morphological Traits in RILs Population Derived from Interspecific Crosses between Pisum sativum × P. fulvum
Previous Article in Special Issue
Deciphering the Mechanism of Wogonin, a Natural Flavonoid, on the Proliferation of Pulmonary Arterial Smooth Muscle Cells by Integrating Network Pharmacology and In Vitro Validation
 
 
Search for Articles:
Title / Keyword
Author / Affiliation / Email
Journal
Article Type
 
 
Advanced Search
 
Section
Special Issue
Volume
Issue
Number
Page
 
Journals
CIMB
Volume 45
Issue 1
10.3390/cimb45010045
cimb-logo
Submit to this Journal Review for this Journal Propose a Special Issue
► Article Menu

Article Menu

share Share announcement Help format_quote Cite question_answer Discuss in SciProfiles
first_page
settings
Order Article Reprints
Font Type:
Arial Georgia Verdana
Font Size:
Aa Aa Aa
Line Spacing:
Column Width:
Background:
Article

In Vitro Antiviral Evaluations of Coldmix®: An Essential Oil Blend against SARS-CoV-2

by
Kemal Hüsnü Can Başer
1,2,
Ayşe Esra Karadağ
3,4,
Sevde Nur Biltekin
5,6,
Murat Ertürk
7 and
Fatih Demirci
2,8,9,*
1
Department of Pharmacognosy, Faculty of Pharmacy, Near East University, N. Cyprus, Mersin 10, 99138 Nicosia, Türkiye
2
Badebio Biotechnology Ltd., ATAP, Anadolu University, Tepebaşı, 26470 Eskişehir, Türkiye
3
Department of Pharmacognosy, School of Pharmacy, Istanbul Medipol University, 34810 Istanbul, Türkiye
4
Department of Pharmacognosy, Graduate School of Health Sciences, Anadolu University, 26470 Eskişehir, Türkiye
5
Department of Pharmaceutical Microbiology, School of Pharmacy, Istanbul Medipol University, 34810 Istanbul, Türkiye
6
Department of Molecular Biology and Genetics, Faculty of Science, Istanbul University, 34134 Istanbul, Türkiye
7
Department of Microbiology, Medical School of Yüksek İhtisas University, 06520 Ankara, Türkiye
8
Department of Pharmacognosy, Faculty of Pharmacy, Anadolu University, 26470 Eskişehir, Türkiye
9
Faculty of Pharmacy, Eastern Mediterranean University, N. Cyprus, Mersin 10, 99628 Famagusta, Türkiye
*
Author to whom correspondence should be addressed.
Curr. Issues Mol. Biol. 2023, 45(1), 677-684; https://doi.org/10.3390/cimb45010045
Submission received: 9 December 2022 / Revised: 30 December 2022 / Accepted: 3 January 2023 / Published: 11 January 2023
(This article belongs to the Collection Application of Natural and Pseudo Natural Products in Drug Discovery and Development)
Browse Figure
Versions Notes

Abstract

:
Coldmix® is a commercially available Eucalyptus aetheroleum and, Abies aetheroleum blend for medicinal applications. In this present study, the in vitro antiviral potential of Coldmix®, and its major constituents 1,8-cineole and α-pinene were evaluated by using the in vitro ACE2 enzyme inhibition assay as well as the direct contact test against SARS-CoV-2. The observed ACE2 enzyme inhibitory activity of Coldmix®, 1,8-cineole, and α-pinene were 72%, 88%, and 80%, respectively; whereas in the direct contact test in the vapor phase, the destruction of the virus was 79.9% within 5 min and 93.2% in the 30th min, respectively. In a similar Coldmix® vapor phase setup using the in vitro cytotoxicity cell assay, E6 VERO healthy cells were experimentally not affected by toxicity. According to the promising initial antiviral results of Coldmix® and the individually tested constituents, detailed further in vivo evaluation using different virus classes is suggested.

1. Introduction

Traditionally, Eucalyptus and Abies essential oils are used specifically to treat respiratory complaints and disorders. It is well documented that Eucalyptus essential oils and preparations are also used for the treatment of pharyngitis, bronchitis, and sinusitis [1,2,3,4,5]. In previous studies, results showing the antimicrobial and antiviral effects of Eucalyptus essential oil were reported [6,7]. It is well documented that Eucalyptus essential oils are prominent with their antimicrobial activities, especially against upper respiratory tract pathogens and infections [8,9,10,11,12]. The Eucalyptus oil was previously demonstrated to be effective against influenza and herpes viruses [13,14]. In addition, the antimicrobial effect of Eucalyptus essential oil combinations was found to be effective, in particular against upper respiratory tract diseases [15]. Both the Eucalyptus oil and its active component 1,8-cineole was observed for its muscle relaxant effect by reducing the smooth muscle contractions in respiratory disorders [16,17,18]. It was also reported that 1,8-cineole was effective against influenza A (H1N1), and inactivated the free influenza A virus by disruption of the envelope structures in vitro [15]. A previous study demonstrated the efficacy of 1,8-cineole in numerous clinical studies conducted in patients with acute and chronic respiratory conditions, including rhinosinusitis, bronchitis, COPD and asthma [18,19].
The inhalation of Abies species preparations showed effective results, especially in microbial and viral infections of the respiratory tract [20,21]. Abies species were inhibitory against upper respiratory pathogenic bacteria, and this effect was observed to increase even more in Abies species rich in α-pinene content [22,23,24,25,26]. The monoterpenes α-pinene as well as 1,8-cineole were previously reported for their in vitro antiviral activity [27]. 1,8-Cineole was recently demonstrated in vitro as an ACE2 inhibitor [28]. ACE2 was one of the receptors causing SARS-CoV-2 [29]. Previous studies reported also that ACE2 is one of the essential receptors for various coronavirus types for cell entry [30,31].
Recent in silico studies showed that essential oils may be effective against COVID-19 by various volatile components [31,32]. The potential effects of Eucalyptus oil and its major constituent 1,8-cineole were recently evaluated against SARS-CoV-2 using in vitro tests and molecular docking techniques [33,34], which suggested the detailed experimentation of volatile compounds and essential oils against coronaviruses.
Effective measures and approaches, such as the application of small-molecule inhibitors and vaccines are investigated to reduce SARS-CoV-2 transmission [35,36]; however, effective and promising drugs still do not exist [37]. As an indispensable resource, traditional medicines [38] and bioactive natural products [39] demonstrated a potential value in countering SARS-CoV-2 infection. In this present study, the in vitro antiviral activity and ACE2 enzyme inhibitory potential of the commercial medical device Coldmix®, and its major constituents 1,8-cineole and α-pinene were studied, to the best of our knowledge for the first time.

2. Materials and Methods

2.1. Materials

If not stated otherwise, all chemicals such as 1,8-cineole, α-pinene, and DMSO were acquired from Sigma/Aldrich at analytical or pharmaceutical grade; the enzyme kits, standard materials and Coldmix® were provided by ENAFARMA Ltd., Istanbul, Turkey.
GC-FID and GC/MS analysis details of Coldmix®: α-pinene: 13.7 ± 0.9%, and 1,8-cineole: 30.7 ± 0.8%.

2.2. ACE2 Enzyme Inhibitory Activity

The test substances were initially dissolved in DMSO < 1% (v/v). The enzyme inhibition experiment was carried out in accordance with the manufacturer’s instructions for the kit “Angiotensin II Converting Enzyme (ACE2) Inhibitor Screening Kit (BioVision, K310 (Waltham, MA, USA))” and the enzyme inhibition of the substances was measured with Ex/Em = 320/420 nm wavelength in a multimode microplate reader (SpectraMax i3 (Sunnyvale, CA, USA)) at fluorescence mode. % inhibition values were calculated for all samples resulting from duplicate data as previously reported [28,40]. Results were illustrated in Figure 1.

2.3. In Vitro Antiviral Activity

The antiviral effectiveness of Coldmix® was tested against SARS-CoV-2 virus (Clinical isolate, GenBank No: MT955161.1) using the Biosafety Level 3 (BSL3) facility at the Antimikrop Ar-Ge Biyosidal Merkezi (Ankara, Turkey), a research center with ISO/IEC 17025 accreditation, which is also licensed by the Turkish Ministry of Health as a competent laboratory for biocidal testing. Virus stock culture was prepared using the Vero E6 cell line, propagated using DMEM-10 (Dulbecco’s Minimum Essential Medium) supplemented with 10% FBS (fetal bovine serum) and pen/strep/fungizone.
The virus suspension was spotted onto the 2 cm2 area on the stainless steel disc, as described in TS EN 16777: 2019 standard, and left to dry in the biosafety cabinet [TS EN 16777:2018: Chemical Disinfectants and Antiseptics—Quantitative Non-Porous Surface Test without Mechanical Action for the Evaluation of Virucidal Activity of Chemical Disinfectants Used in the Medical Area—Test Method and Requirements (Phase 2/Step 2) European Committee for Standardization; Brussels, Belgium: 2019 2019)]. After the discs were dried, 4 control discs were left in the biosafety cabinet, while 6 test discs were placed in a 15 L plastic container with a lid. A piece of cloth fabric onto which 5–6 drops of Coldmix® applied was also placed into one corner of the box, making sure that the virus containing discs were about 20 cm away from the cloth. After tightly closing the box with its lid, it was placed into an incubator for 5 and 30 min at 30 ± 1 °C. The control disks were kept in a sealed petri dish at the lower shelves of the incubator. After the exposure times, disks were individually processed as described in the EN 16777: 2019 standard, by performing an extraction procedure which essentially involves vortexing the discs for 30 s in 0.9 mL complete DMEM. The harvest of each disc was then serially diluted in 10-folds using complete DMEM, and transferred into a 96-well tissue culture plate containing Vero E6 cell suspension at 20,000 cell/well. The plates were incubated at 37 ± 10 °C, 5% CO2 environment for 4 days [41,42,43]. The 50% tissue culture infective dose (TCID50) of the tests and controls were calculated using the Spearman-Kärber method, as described by EN 16777: 2019 standard. The titers were expressed as log10 TCID50.

2.4. Cytotoxicity

To measure the extent of the cellular toxicity, the cells were directly exposed to the vapor of Coldmix® in a closed chamber. For this, E6 VERO cell suspension was seeded into 96-well plates at 20,000 cells/well and incubated in a 37 °C 5% CO2 incubator overnight. The next day, a water wetted tissue paper was placed in a 15 L plastic container with a lid, and kept in an incubator at 37 °C and 90% humidity. 10 × 10 cm fabric was placed inside the container by folding it in half in a corner. 6–8 drops of Coldmix® were dropped on the fabric, and the lid of the box was closed and placed into 37 °C incubator for vapor conditioning inside the container. After five minutes, the contents of the cell plates was replaced with 0.025 mL/well of DMEM containing 2% FBS to prevent drying of cell layer during the incubation period. Subsequently, half of the plates were covered with an adhesive plate cover to avoid exposure to Coldmix® vapor. The closed part was used as a control. The plates were then placed into the container conditioned as described above. After the plates were placed in the container, one plate was removed after 5 min, and the other 30 min, followed by MTT staining. For this, MTT [(4,5-dimethyl-thiazoyl)-2,5-diphenyl-SH-tetrazolium bromide] dye solution at 0.5 mg/mL concentration in DMEM containing 2% FBS added into all wells. After 3 h of incubation at 37 °C, and 5% CO2, the content was replaced with DMSO to extract the formazan crystals. Subsequently, the optical density (OD) was read on a microplate reader (Tecan Sunrise, Mannedorf, Zurich, Switzerland) at 570 nm. For the calculation of the results, the mean values for the cells exposed to Coldmix® vapor and untreated (control) for 5 and 30 min were recorded.

2.5. Statistical Analysis

In vitro data was expressed as mean ± standard error of the mean (Mean ± SEM). The statistical significance between groups was analysed by One-way ANOVA (followed by Dunnett’s post hoc test) and Paired Samples T-Test. The statistical analysis was carried out using the GraphPad Prism 7.0 and SPSS Statistics 22.0 software. The p < 0.05 was considered as statistically significant.

3. Results and Discussion

3.1. ACE2 Enzyme Inhibitory Activity

In the present study, the in vitro ACE2 enzyme inhibitory activity of the essential oil blend Coldmix® (at 20 µg/mL concentration), and its major components 1,8-cineole as well as α-pinene (at 5 µg/mL concentration) were evaluated, respectively. α-Pinene showed a relatively higher ACE2 inhibitory activity compared to 1,8-cineole, and the Coldmix® itself. While the Coldmix® showed 72% ACE2 enzyme inhibitory activity, the major components 1,8-cineole and α-pinene showed 88.12%, and 80.03% enzyme inhibition, respectively, as shown in Figure 1.
The possible utilization and application of ACE2 inhibitors against COVID-19 infections were reported in previous studies [44]. As is well known, the interaction of the COVID-19 virus spike and ACE2 is necessary for virus infection; any agent that interrupts its interaction, the human monoclonal antibody based on the receptor binding domain, and the recombinant human ACE2 protein (rhuACE2) are current targets [45]. The ACE2 inhibitory activity evaluation of 1,8-cineole of the Rosmarinus essential oil was previously reported [28] also in correlation with the findings in this present study. In addition, in recent studies and reports [40] of our group on in vitro ACE2 enzyme inhibition of Eucalyptus essential oil was also relatively high, suggesting that the recent effect of Coldmix® may be influenced by Eucalyptus oil. As Coldmix® inhibited in vitro the ACE2 system, the commercial product and its major components could be effective against SARS-CoV-2. This was also supported by the in vitro antiviral effects, which is also reported in this work for the first time.

3.2. Antiviral Activity and Cytotoxicity of Vapor Phase

In this present work, the in vitro antiviral activity of the Coldmix® was tested against the human pathogenic SARS-CoV-2 virus, where the initial results are shown in Table 1. The essential oil blend Coldmix® indicated remarkable antiviral activity against the tested human pathogenic virus at the tested concentration as low as 0.1 mg/mL in the contact range of 5 to 30 min. Visually confirmed, Coldmix® destroyed the virus by 79.89% in the 5th min, and 93.23% in the 30th min, as per the experimental measurements.
1,8-Cineole, a component of Coldmix®, is known for its antiviral potential against different human pathogen DNA and RNA viruses such as rhinoviruses, HSV-1, hepatitis A virus, herpes viruses (HSV), adenoviruses (ADV), hepatitis B virus, coxsackievirus B1 (CVB1), and enterovirus 71 (EV71) [46,47,48,49]. As a result of this in vitro study, it was observed that the product, which is rich in 1,8-cineole, is relatively effective against the coronavirus tested. It can be proposed that 1,8-cineole may contribute or is even responsible for this effect. According to previous studies, α-pinene, was tested against different human pathogenic viruses, and promising results were observed [47]. In addition, a study on mixtures of aroma substances, volatile compounds or essential oils showed that in some cases, individual volatile components showed relatively lower effects compared to their initial essential oils, respectively [50]. The medical device Coldmix®, is also such a unique combination of volatile components in this respect, which may have future potential in natural antiviral applications on clinical experiments.
Based to the experimental data, the tested Coldmix® product was also found to be relatively safe in vitro according to the cytotoxicity data on healthy Vero cells in the vapor phase for 5 min and 30 min, using a similar setup to the antiviral assay as explained in detail. As Coldmix® is generally used by inhalation, its cytotoxicity was elaborated in the vapor phase. After 5 min and 30 min of exposure, no difference was observed between the negative control group and the Coldmix® test group, which suggests the relative safe use of this special essential oil blend in the vapor phase.
The COVID-19 pandemic is currently still an important and definitive health problem in the world. Essential oils are known to have a broad bioactivity spectrum such as antimicrobials and also antivirals since ancient times. There are numerous studies that reveal the potential antiviral effects of essential oils on different viruses such as polio type 1, ECHO 9, Coxsackie B1, adeno type 2, herpes simplex (HSV) type 1 and 2, and H1N1 alone or in combination [15,50,51]. Some of these viruses belong to the coronaviruses, and even though limited, there are antiviral effect studies of coronaviruses on essential oils [28,40,41,52]. The effectiveness of the proprietary commercial essential oil blend Coldmix®, evaluated against coronavirus, both by direct contact with the virus and by ACE2 enzyme inhibition, was demonstrated in this study for the first time. The antiviral potential of other essential oils components and volatile metabolites remain as a potential for future detailed studies.

4. Conclusions

The proprietary medicinal product Coldmix®, which has been used for more than a decade, is a special blend of essential oils containing volatile compounds which may be used both for the prevention and treatment against coronaviruses, among other microorganisms. However, further detailed bioassays as well as clinical studies are needed to confirm the efficacy against SARS-CoV-2.

Author Contributions

Conceptualization K.H.C.B. and F.D.; methodology, A.E.K., S.N.B. and M.E.; software, validation, A.E.K. and S.N.B.; formal analysis, A.E.K., M.E. and S.N.B.; investigation, S.N.B. and A.E.K.; resources, F.D. and K.H.C.B.; data curation, S.N.B. and M.E.; writing—original draft preparation, K.H.C.B., F.D. and A.E.K.; writing—review and editing, F.D., K.H.C.B. and A.E.K.; visualization, S.N.B. All authors have read and agreed to the published version of the manuscript.

Funding

This research was supported financially by EnaFarma Ltd.

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

All data are available in this article.

Acknowledgments

The “antiviral use of Coldmix®” was subject for a patent file application TPE 2021/006518. This work is dedicated to the 100th anniversary of Republic of Türkiye.

Conflicts of Interest

The authors declare no conflict of interest.

List of Abbreviations

ACE2: Angiotensin Convertying Enzyme-2, ADV: Adenoviruses, BSL3: Biosafety Level 3, COPD: Chronic obstructive pulmonary disease, COVID-19: Coronavirus disease-19, CVB: coxsackievirus B1, DMEM: Dulbecco’s Minimum Essential Medium, DMSO: Dimetyhlsulfoxide, EV71: enterovirus 71, FBS: Fetal Bovine Serum, HSV: Herpes Virus, MTT: (4,5-dimethyl-thiazoyl)-2,5-diphenyl-SH-tetrazolium bromide.

References

  1. Ghnaya, A.B.; Hanana, M.; Kaderi, M.; Amri, I.; Hamrouni, L. Eucalyptus erythrocorys L. notes ethnobotanique et phytopharmacologique. Phytotherapie 2015, 13, 262–266. [Google Scholar] [CrossRef]
  2. Paniagua-Zambrana, N.Y.; Bussmann, R.W.; Romero, C. Eucalyptus citriodora Hook. Eucalyptus globulus Labill. Myrtaceae. In Ethnobotany of the Andes; Springer: Cham, Switzerland, 2020; pp. 829–836. [Google Scholar]
  3. Orch, H.; Zidane, L.; Douira, A. Ethnobotanical study of plants used in the treatment of respiratory diseases in a population bordering the forest of Izarène. J. Pharm. Pharmacogn. Res. 2020, 8, 392–409. [Google Scholar]
  4. Menković, N.; Šavikin, K.; Tasić, S.; Zdunić, G.; Stešević, D.; Milosavljević, S.; Vincek, D. Ethnobotanical study on traditional uses of wild medicinal plants in Prokletije Mountains (Montenegro). J. Ethnopharmacol. 2011, 133, 97–107. [Google Scholar] [CrossRef] [PubMed]
  5. Bulut, G.; Tuzlacı, E.; Tuzlaci, E. An Ethnobotanical Study of Medicinal Plants in Bayramiç (Çanakkale-Turkey). Marmara Pharm. J. 2015, 19, 268–282. [Google Scholar] [CrossRef]
  6. Elaissi, A.; Rouis, Z.; Salem, N.A.B.; Mabrouk, S.; ben Salem, Y.; Salah, K.B.H.; Aouni, M.; Farhat, F.; Chemli, R.; Harzallah-Skhiri, F.; et al. Chemical composition of 8 eucalyptus species’ essential oils and the evaluation of their antibacterial, antifungal and antiviral activities. BMC Complement. Altern. Med. 2012, 12, 81. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  7. Cermelli, C.; Fabio, A.; Fabio, G.; Quaglio, P. Effect of eucalyptus essential oil on respiratory bacteria and viruses. Curr. Microbiol. 2008, 56, 89–92. [Google Scholar] [CrossRef]
  8. Damjanović-Vratnica, B.; Đakov, T.; Šuković, D.; Damjanović, J. Antimicrobial Effect of Essential Oil Isolated from Eucalyptus globulus Labill. from Montenegro. Czech J. Food Sci. 2011, 29, 277–284. [Google Scholar] [CrossRef] [Green Version]
  9. Hafsa, J.; ali Smach, M.; Ben Khedher, M.R.; Charfeddine, B.; Limem, K.; Majdoub, H.; Rouatbi, S. Physical, antioxidant and antimicrobial properties of chitosan films containing Eucalyptus globulus essential oil. LWT-Food Sci. Technol. 2016, 68, 356–364. [Google Scholar] [CrossRef]
  10. Sartorelli, P.; Marquioreto, A.D.; Amaral-Baroli, A.; Lima, M.E.L.; Moreno, P.R.H. Chemical composition and antimicrobial activity of the essential oils from two species of Eucalyptus. Phyther. Res. 2007, 21, 231–233. [Google Scholar] [CrossRef]
  11. Gilles, M.; Zhao, J.; An, M.; Agboola, S. Chemical composition and antimicrobial properties of essential oils of three Australian Eucalyptus species. Food Chem. 2010, 119, 731–737. [Google Scholar] [CrossRef]
  12. Delaquis, P.J.; Stanich, K.; Girard, B.; Mazza, G. Antimicrobial activity of individual and mixed fractions of dill, cilantro, coriander and Eucalyptus essential oils. Int. J. Food Microbiol. 2002, 74, 101–109. [Google Scholar] [CrossRef]
  13. Schnitzler, P.; Schön, K.; Reichling, J. Antiviral activity of Australian tea tree oil and eucalyptus oil against Herpes simplex virus in cell culture. Pharmazie 2001, 56, 343–347. [Google Scholar]
  14. Usachev, E.V.; Pyankov, O.V.; Usacheva, O.V.; Agranovski, I.E. Antiviral activity of tea tree and eucalyptus oil aerosol and vapour. J. Aerosol Sci. 2013, 59, 22–30. [Google Scholar] [CrossRef]
  15. Brochot, A.; Guilbot, A.; Haddioui, L.; Roques, C. Antibacterial, antifungal, and antiviral effects of three essential oil blends. Microbiologyopen 2017, 6, 459. [Google Scholar] [CrossRef]
  16. Coelho-De-Souza, L.N.; Leal-Cardoso, J.H.; De Abreu Matos, F.J.; Lahlou, S.; Magalhães, P.J.C. Relaxant effects of the essential oil of Eucalyptus tereticornis and its main constituent 1,8-cineole on guinea-pig tracheal smooth muscle. Planta Med. 2005, 71, 1173–1175. [Google Scholar] [CrossRef]
  17. Bastos, V.P.; Brito, T.S.; Lima, F.J.; Pinho, J.P.; Lahlou, S.; Abreu Matos, F.J.; Santos, A.A.; Caldas Magalhães, P.J. Inhibitory effect of 1,8-cineole on guinea-pig airway challenged with ovalbumin involves a preferential action on electromechanical coupling. Clin. Exp. Pharmacol. Physiol. 2009, 36, 1120–1126. [Google Scholar] [CrossRef]
  18. Juergens, U.R.; Dethlefsen, U.; Steinkamp, G.; Gillissen, A.; Repges, R.; Vetter, H. Anti-inflammatory activity of 1.8-cineole (eucalyptol) in bronchial asthma: A double-blind placebo-controlled trial. Respir. Med. 2003, 97, 250–256. [Google Scholar] [CrossRef] [Green Version]
  19. Juergens, L.J.; Worth, H.; Juergens, U.R. New perspectives for mucolytic, anti-inflammatory and adjunctive therapy with 1,8-cineole in COPD and asthma: Review on the new therapeutic approach. Adv. Ther. 2020, 37, 1737–1753. [Google Scholar] [CrossRef] [Green Version]
  20. Böhme, K.; Barros-Velázquez, J.; Calo-Mata, P.; Aubourg, S.P. Antibacterial, antiviral and antifungal activity of essential oils: Mechanisms and applications. In Antimicrobial Compounds: Current Strategies and New Alternatives; Springer: Berlin/Heidelberg, Germany, 2014; pp. 51–81. [Google Scholar]
  21. Lee, S.Y.; Kirn, S.H.; Park, M.J.; Lee, S.S.; Choi, I.G. Antibacterial activity of essential oil from Abies holophylla against respiratory tract bacteria. J. Korean Wood Sci. Technol. 2014, 42, 533–542. [Google Scholar] [CrossRef] [Green Version]
  22. Baran, S.; von Reuss, S.H.; König, W.A.; Kalemba, D. Composition of the Essential oil of Abies koreana Wils. Flavour Fragr. J. 2007, 22, 78–83. [Google Scholar] [CrossRef]
  23. Pichette, A.; Larouche, P.L.; Lebrun, M.; Legault, J. Composition and antibacterial activity of Abies balsamea essential oil. Phyther. Res. 2006, 20, 371–373. [Google Scholar] [CrossRef] [PubMed]
  24. Lee, J.H.; Hong, S.K. Comparative analysis of chemical compositions and antimicrobial activities of Essential oils from Abies holophylla and Abies koreana. J. Microbiol. Biotechnol. 2009, 19, 372–377. [Google Scholar] [CrossRef] [PubMed]
  25. Kartnig, T.; Still, F.; Reinthaler, F. Antimicrobial activity of the essential oil of young pine shoots (Picea abies L.). J. Ethnopharmacol. 1991, 35, 155–157. [Google Scholar] [CrossRef] [PubMed]
  26. Bagci, E.; Digrak, M. Antimicrobial Activity of Essential Oils of some Abies (Fir) Species from Turkey. Flavour Fragr. J. 1996, 11, 251–256. [Google Scholar] [CrossRef]
  27. Loizzo, M.R.; Saab, A.M.; Tundis, R.; Statti, G.A.; Menichimi, F.; Lampronti, D.; Gambari, R.; Cinatl, J.; Doerr, H.W. Phytochemical analysis and in vitro antiviral activities of the essential oils of seven Lebanon species. Chem. Biodivers. 2008, 5, 461–470. [Google Scholar] [CrossRef]
  28. Demirci, F.; Karadağ, A.E.; Biltekin, S.N.; Demirci, B. In vitro ACE2 and 5-LOX Inhibition of Rosmarinus officinalis L. Essential Oil and its Major Component 1,8-Cineole. Rec. Nat. Prod. 2022, 16, 194–199. [Google Scholar]
  29. Amirfakhryan, H.; Safari, F. Outbreak of SARS-CoV2: Pathogenesis of infection and cardiovascular involvement. Hell. J. Cardiol. 2021, 62, 13–23. [Google Scholar] [CrossRef]
  30. Kuba, K.; Imai, Y.; Rao, S.; Gao, H.; Guo, F.; Guan, B.; Huan, Y.; Yang, P.; Zhang, Y.; Deng, W.; et al. A crucial role of angiotensin converting enzyme 2 (ACE2) in SARS coronavirus–induced lung injury. Nat. Med. 2005, 11, 875–879. [Google Scholar] [CrossRef]
  31. Li, W.; Moore, M.J.; Vasilieva, N.; Sui, J.; Wong, S.K.; Berne, M.A.; Somasundaran, M.; Sullivan, J.L.; Luzuriaga, K.; Greenough, T.C.; et al. Angiotensin-converting enzyme 2 is a functional receptor for the SARS coronavirus. Nature 2003, 426, 450–454. [Google Scholar] [CrossRef] [Green Version]
  32. Pillaiyar, T.; Flury, P.; Krüger, N.; Su, H.; Schäkel, L.; Barbosa Da Silva, E.; Laufer, S.A. Small-Molecule Thioesters as SARS-CoV-2 Main Protease Inhibitors: Enzyme Inhibition, Structure–Activity Relationships, Antiviral Activity, and X-ray Structure Determination. J. Med. Chem. 2022, 65, 9376–9395. [Google Scholar] [CrossRef]
  33. Da Silva, J.K.R.; Figueiredo, P.L.B.; Byler, K.G.; Setzer, W.N. Essential oils as antiviral agents. Potential of essential oils to treat SARS-CoV-2 infection: An in−silico investigation. Int. J. Mol. Sci. 2020, 21, 3426. [Google Scholar] [CrossRef]
  34. Nadjib Boukhatem, M.; Mohamed Nadjib, B. Effective antiviral activity of essential oils and their characteristic terpenes against coronaviruses: An update. J. Pharmacol. Clin. Toxicol. 2020, 8, 1138. [Google Scholar]
  35. Tomalka, J.A.; Suthar, M.S.; Deeks, S.G.; Sekaly, R.P. Fighting the SARS-CoV-2 pandemic requires a global approach to understanding the heterogeneity of vaccine responses. Nature Immunol. 2022, 23, 360–370. [Google Scholar] [CrossRef]
  36. Wang, Z.; Yang, L. GS-5734: A potentially approved drug by FDA against SARS-CoV-2. New J. Chem. 2020, 44, 12417–12429. [Google Scholar] [CrossRef]
  37. Wang, Z.; Yang, L. Chinese herbal medicine: Fighting SARS-CoV-2 infection on all fronts. J. Ethnopharmacol. 2020, 270, 113869. [Google Scholar] [CrossRef]
  38. Yang, L.; Wang, Z. Natural products, alone or in combination with FDA-approved drugs, to treat COVID-19 and lung cancer. Biomedicines 2021, 9, 689. [Google Scholar] [CrossRef]
  39. Asif, M.; Saleem, M.; Saadullah, M.; Yaseen, H.S.; Al Zarzour, R. COVID-19 and therapy with essential oils having antiviral, anti-inflammatory, and immunomodulatory properties. Inflammopharmacology 2020, 28, 1153–1161. [Google Scholar] [CrossRef]
  40. Ak Sakallı, E.; Teralı, K.; Karadağ, A.E.; Biltekin, S.N.; Koşar, M.; Demirci, B.; Demirci, F. In vitro and in silico evaluation of ACE2 and LOX inhibitory activity of Eucalyptus essential oils, 1,8-cineole, and citronellal. Nat. Prod. Com. 2022, 17, 1934578X221109409. [Google Scholar] [CrossRef]
  41. Biltekin, S.N.; Karadağ, A.E.; Demirci, B.; Demirci, F. ACE2 and LOX Enzyme Inhibitions of Different Lavender Essential Oils and Major Components Linalool and Camphor. ACS Omega 2022, 7, 36561–36566. [Google Scholar] [CrossRef]
  42. Yao, X.; Ye, F.; Zhang, M.; Cui, C.; Huang, B.; Niu, P.; Liu, X.; Zhao, L.; Dong, E.; Song, C.; et al. In vitro antiviral activity and projection of optimized dosing design of hydroxychloroquine for the treatment of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Clin. Infect. Dis. 2020, 71, 732–739. [Google Scholar] [CrossRef] [Green Version]
  43. Şakalar, Ç.; Ertürk, M. Inactivation of airborne SARS-CoV-2 by thyme volatile oil vapor phase. J. Virol. Methods 2022, 24, 114660. [Google Scholar] [CrossRef] [PubMed]
  44. Liu, M.; Wang, T.; Zhou, Y.; Zhao, Y.; Zhang, Y.; Li, J. Potential role of ACE2 in coronavirus disease 2019 (COVID-19) prevention and management. J. Transl. Intern. Med. 2020, 8, 9–19. [Google Scholar] [CrossRef] [PubMed]
  45. Leng, Z.; Zhu, R.; Hou, W.; Feng, Y.; Yang, Y.; Han, Q.; Shan, G.; Meng, F.; Du, D.; Wang, S.; et al. Transplantation of ACE2-mesenchymal stem cells improves the outcome of patients with COVID-19 pneumonia. Aging Dis. 2020, 11, 216–228. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  46. Müller, J.; Greiner, J.F.W.; Zeuner, M.; Brotzmann, V.; Schäfermann, J.; Wieters, F.; Widera, D.; Sudhoff, H.; Kaltschmidt, B.; Kaltschmidt, C. 1,8-Cineole potentiates IRF3-mediated antiviral response in human stem cells and in an ex vivo model of rhinosinusitis. Clin. Sci. 2016, 130, 1339–1352. [Google Scholar] [CrossRef] [PubMed]
  47. Astani, A.; Reichling, J.; Schnitzler, P. Comparative study on the antiviral activity of selected monoterpenes derived from essential oils. Phyther. Res. 2010, 24, 673–679. [Google Scholar] [CrossRef]
  48. Battistini, R.; Rossini, I.; Ercolini, C.; Goria, M.; Callipo, M.R.; Maurella, C.; Pavoni, E.; Serracca, L. Antiviral activity of essential oils against hepatitis a virus in soft fruits. Food Environ. Virol. 2019, 11, 90–95. [Google Scholar] [CrossRef]
  49. Chiang, L.C.; Ng, L.T.; Cheng, P.W.; Chiang, W.; Lin, C.C. Antiviral activities of extracts and selected pure constituents of Ocimum basilicum. Clin. Exp. Pharmacol. Physiol. 2005, 32, 811–816. [Google Scholar] [CrossRef]
  50. Sivropoulou, A.; Nikolaou, C.; Papanikolaou, E.; Kokkini, S.; Lanaras, T.; Arsenakis, M. Antimicrobial, cytotoxic, and antiviral activities of Salvia fruticosa essential oil. J. Agric. Food Chem. 1997, 45, 3197–3201. [Google Scholar] [CrossRef]
  51. Garozzo, A.; Timpanaro, R.; Bisignano, B.; Furneri, P.M.; Bisignano, G.; Castro, A. In vitro antiviral activity of Melaleuca alternifolia essential oil. Lett. Appl. Microbiol. 2009, 49, 806–808. [Google Scholar] [CrossRef]
  52. Catella, C.; Camero, M.; Lucente, M.S.; Fracchiolla, G.; Sblano, S.; Tempesta, M.; Lanave, G. Virucidal and antiviral effects of Thymus vulgaris essential oil on feline coronavirus. Res. Vet. Sci. 2021, 137, 44–47. [Google Scholar] [CrossRef]
Cimb 45 00045 g001 550
Figure 1. The in vitro ACE2 inhibition graph of Coldmix® and major components, presenting the mean ± SD values (n = 6). * p < 0.05 *** p < 0.001 determined by one-way ANOVA using Dunnet’s multiple comparison test.
Figure 1. The in vitro ACE2 inhibition graph of Coldmix® and major components, presenting the mean ± SD values (n = 6). * p < 0.05 *** p < 0.001 determined by one-way ANOVA using Dunnet’s multiple comparison test.
Cimb 45 00045 g001
Table
Table 1. The median tissue culture infective dose (TCID50) of Coldmix® and test samples against SARS-CoV-2.
Table 1. The median tissue culture infective dose (TCID50) of Coldmix® and test samples against SARS-CoV-2.
TESTTest MaterialLog TCID50Mean
(±)		Results
Virus TitrationStock Virus7.5--
Virucidal Test
(5 min)		Control D 16.176.25R = Log K − Log T
R = 0.69		72.45%
Control D 26.33
Coldmix® Test D 15.835.53
Coldmix® Test D 25.50
Coldmix® Test D 35.33
Virucidal Test
(30 min)		Control D 16.176.17R = Log K − Log T
R = 1.17		92.41%
Control D 26.17
Coldmix® Test D 14.835.00
Coldmix® Test D 25.17
Coldmix® Test D 35.00
Disclaimer/Publisher’s Note: The statements, opinions and data contained in all publications are solely those of the individual author(s) and contributor(s) and not of MDPI and/or the editor(s). MDPI and/or the editor(s) disclaim responsibility for any injury to people or property resulting from any ideas, methods, instructions or products referred to in the content.

Share and Cite

MDPI and ACS Style

Başer, K.H.C.; Karadağ, A.E.; Biltekin, S.N.; Ertürk, M.; Demirci, F. In Vitro Antiviral Evaluations of Coldmix®: An Essential Oil Blend against SARS-CoV-2. Curr. Issues Mol. Biol. 2023, 45, 677-684. https://doi.org/10.3390/cimb45010045

AMA Style

Başer KHC, Karadağ AE, Biltekin SN, Ertürk M, Demirci F. In Vitro Antiviral Evaluations of Coldmix®: An Essential Oil Blend against SARS-CoV-2. Current Issues in Molecular Biology. 2023; 45(1):677-684. https://doi.org/10.3390/cimb45010045

Chicago/Turabian Style

Başer, Kemal Hüsnü Can, Ayşe Esra Karadağ, Sevde Nur Biltekin, Murat Ertürk, and Fatih Demirci. 2023. "In Vitro Antiviral Evaluations of Coldmix®: An Essential Oil Blend against SARS-CoV-2" Current Issues in Molecular Biology 45, no. 1: 677-684. https://doi.org/10.3390/cimb45010045

APA Style

Başer, K. H. C., Karadağ, A. E., Biltekin, S. N., Ertürk, M., & Demirci, F. (2023). In Vitro Antiviral Evaluations of Coldmix®: An Essential Oil Blend against SARS-CoV-2. Current Issues in Molecular Biology, 45(1), 677-684. https://doi.org/10.3390/cimb45010045

Article Metrics

Back to TopTop