Escherichia coli에 대한 17가지 에센셜 오일의 항균 활성

The Antibacterial Activity of 17 Essential Oils against Escherichia coli

17种精油对大肠杆菌的抗菌活性

Article information

Asian J Beauty Cosmetol. 2022;20(1):33-41
Publication date (electronic) : 2022 March 23
doi : https://doi.org/10.20402/ajbc.2021.0263
1Department of Public Health Science, Dankook University, Cheonan-si, Chungchengnam-do, Korea
2Department of Clinical Medical Science, Dankook University, Cheonan-si, Chungchengnam-do, Korea
3Department of Biomedical Laboratory Science, Dankook University, Cheonan-si, Chungchengnam-do, Korea
박정민1, 육영삼2, 이영기3, 김재경3,
1단국대학교 보건학과, 충청남도 천안시, 한국
2단국대학교 임상의과학과, 충청남도 천안시, 한국
3단국대학교 임상병리학과, 충청남도 천안시, 한국
*Corresponding author: Jae Kyung Kim, Department of Biomedical Laboratory Science, Dankook University, 119 Dandae-ro, Dongnam-gu, Cheonan-si, Chungchengnam-do 31116, Korea

Jung Min Park and Young Sam Yuk contributed equally to this work.

Received 2021 November 24; Revised 2021 December 14; Accepted 2022 February 10.

Abstract

목적

본 연구의 목적은 17가지 에센셜 오일(EO)이 사용 농도 및 추출 부위에 따른 항균 활성을 포함하여 Escherichia coli (E. coli)에 미치는 영향을 조사하는 것입니다.

방법

모든 17개의 EO는 0.5% (v/v) 및 1% (v/v) 농도에서 단계적 희석에 의해 제조되었습니다. EO을 E. coli 배양액과 함께 96-well plates에 분배하고 18시간 동안 인큐베이션 했습니다. 그런 다음 마이크로플레이트 판독기를 사용하여 흡광도를 측정했습니다.

결과

배양 흡광도는 0.5% (v/v) 및 1% (v/v) 농도에서 타임 화이트(TM), 팔마로사(PR) 및 로즈마리 버베논(RM)이 암피실린과 유사한 높은 항균 활성을 나타냄을 보여주었습니다. 멜리사 트루(MS), RM, PR, 라벤더 불가리안(LV), 레몬(LM), 페퍼민트 프리미엄(PM), 유칼립투스 블루검(EC)도 농도 의존적으로 항균 활성이 증가한 것으로 나타났다. 마지막으로, 잎에서 추출한 TM, MS, RM, PR 및 레몬그라스(LG)는 다른 기관에서 추출한 추출물보다 더 높은 항균 활성을 보였다.

결론

잎에서 추출한 TM은 우수한 항균 효과를 나타냈습니다. 동일한 수준의 암피실린에서 박테리아 성장을 나타냈다. 피부의 염증반응에서 세균의 작용을 억제하는 천연물질로 맞춤형 화장품의 원료로 사용될 수 있습니다.

Trans Abstract

Purpose

The purpose of this study was to investigate the effects of 17 essential oils (EOs) on Escherichia coli (E. coli), including their antibacterial activity according to the concentration used and its extraction site.

Methods

All 17 EOs were prepared by step dilution at concentrations of 0.5% (v/v) and 1% (v/v). EOs were dispensed into 96-well plates with bacterial culture aliquots and incubated for 18 h; then, their absorbance was measured using a microplate reader.

Results

Culture absorbance showed that thyme white (TM), palmarosa (PR) and rosemary verbenone (RM) at concentrations of 0.5% (v/v) and 1% (v/v) showed high antibacterial activity, similar to that of ampicillin. Melisa true (MS), RM, PR, lavender Bulgarian (LV), lemon (LM), peppermint premium (PM) and eucalyptus bluegum (EC) also showed increased antibacterial activity in a concentration-dependent manner. Finally, TM, MS, RM, PR, and lemongrass (LG) extracted from leaves showed higher antibacterial activity than extracts from other organs.

Conclusion

TM extracted from leaves showed an excellent antibacterial effect; it exhibited bacterial growth at the same level of ampicillin. It is a natural substance that suppresses the action of bacteria in the inflammatory reaction of the skin and that may be used as a raw material for customized cosmetics.

Trans Abstract

目的

探讨 17 种精油(EO)对大肠杆菌(Escherichia coli, E. coli)的影响,包括根据使用浓度及其提取部位的抗菌活性。

方法

所有 17种精油均通过分步稀释制备,浓度分别为 0.5% (v/v)和1% (v/v)。将精油分装到96孔板和细菌培养等分试样中并孵育18小时;然后,使用酶标仪测量它们的吸光度。

结果

培养吸光度显示百里香(thyme white, TM)、 玫瑰草(palmarosa, PR)和马鞭草酮迷迭香(rosemary verbenone, RM)在浓度为0.5% (v/v)和1% (v/v)时,显示出与氨苄青霉素相似的高抗菌活性。Melisa true (MS)、RM、PR、保加利亚薰衣草(lavender bulgarian, LV)、柠檬(lemon, LM)、薄荷特级(peppermint premium, PM)和蓝胶尤加利(eucalyptus blue gum, EC)也以浓度依赖性方式显示出提高其抗菌活性。最后,从叶子中提取的TM、MS、RM、PR和柠檬草(lemongrass, LG)显示出比其他器官提取物具有更高的抗菌活性。

结论

从叶子中提取的TM具有优良的抑菌效果;它表现出与氨苄青霉素相同水平的细菌生长。它是一种天然物质,可抑制细菌在皮肤炎症反应中的作用,可用作定制化妆品的原料。

Introduction

Escherichia coli, a gram-negative bacterium, is an opportunistic pathogen in humans (Bondarenko et al., 2018) that is commonly present in the human gut. Though E. coli is generally a harmless symbiotic bacterium (Croxen et al., 2013) it may cause urinary tract infections, gastrointestinal infections, and other local tissue and organ infections under certain conditions (Lee et al., 2020b; Wood, 2009). Essential oils (EOs), used in aromatherapy, are highly concentrated oils (Dunning, 2013) extracted from the flowers, fruits, stems, roots, and resins of plants. Frequently, EOs have various antibacterial, antifungal, antiviral, anti-inflammatory, antioxidant, and insect repellent effects as well as hormonal action (Ali et al., 2015). Research on the antibacterial effects of EOs is being actively conducted in various fields (Jo et al., 2018; Park & Kang, 2020; Wang et al., 2020; Wu et al., 2019), but there are not many comparisons on the antibacterial activity of several types of EOs against E. coli.

In this study, we investigated the antibacterial activity of 17 different EOs, widely used in daily life, against E. coli. The antibacterial activity of each type of EO, at concentrations of 0.5% and 1% (v/v), was tested; in addition, the antibacterial activity of different plant organ extracts was studied. Our data suggest that EOs can be actively used as natural antibacterial materials in related industries (food and cosmetics manufacturing).

Materials and Methods

1. Materials

1) Bacteria

The E. coli strain used in this study (KCTC, 11643) was purchased from the Korean Collection for Type Cultures (KCTC).

2) EOs

In these experiments, 17 of La Selection products from The Certification Academy for Holistic Aromatherapy were used (Table 1).

Essential oils used in this study, and tissue/organ extraction sites

3) Equipment

A FlexStation 3 Multi-Mode Microplate Reader (FlexStation 3; Molecular Devices, USA) was used in the course of these experiments.

2. Methods

1) Culture conditions

E. coli strains were subcultured at 37℃ for 18 h on nutrient agar (NA; Difco™, France), and diluted in nutrient broth (NB) to match 0.5 McFarland medium.

2) Oil treatment

EOs were serially diluted in NB to two final concentrations: 0.5% (v/v) and 1% (v/v).

Bacterial suspensions were dispensed into 96-well plates (100 µL/well), and 100 µL of an EO 0.5% (v/v) or 1% (v/v) EO was also dispensed in each well. The control group had 8 µg/mL ampicillin (Kisan Bio, Korea) added (instead of any EO) according to a sensitivity criterion. The cells were incubated for 18 h at 37℃.

3) Bacterial growth determination

Optical density (OD) was measured at 600 nm as an endpoint using a spectrophotometer absorption program for the FlexStation 3 reader. Accuracy was improved with three replicates.

This study was conducted with the approval of the Ethics Committee of Dankook University (IRB File No. NON2021-002). The study was also conducted in accordance with the tenets of the Declaration of Helsinki.

Results

1. Evaluation of the antibacterial activity of 17 EOs in comparison with an antibiotic

Compared with the effect of ampicillin on E. coli, TM showed the same antibacterial activity and PR and RM showed a concentration-dependent antibacterial activity similar to that of ampicillin (Figure 1).

Figure 1.

Antibacterial activity of different essential oils and ampicillin.

The bars show the antibacterial activity of 17 EOs and ampicillin expressed as the optical density of bacterial cultures. Among the EOs, only TM, PR, and RM showed the same antibacterial activity as ampicillin. The horizontal line marks the results obtained with ampicillin, as a reference. EO, essential oil; TM, thyme white; PR, palmarosa; RM, rosemary verbenone; MS, melisa true; LG, lemongrass; RW, rosewood; LV, lavender Bulgarian; LM, Lemon; CP, Cypress; CW, cedarwood atias; TT, tea tree; BG, bergamot; OR, orange sweet; SW, sandalwood; PC, pachuli; PM, peppermint premium; EC, eucalyptus bluegum.

LM, LV, RW, LG, and MS inhibited E. coli growth, mainly at 1% concentrations, but were less effective than ampicillin. All other EOs showed no antibacterial effect; some might even stimulate bacterial growth.

2.Evaluation of the antibacterial activity of 17 EOs at different concentrations

After comparing the activities of the EOs tested at concentrations of 0.5% (v/v) and 1% (v/v), it was observed that TM had the highest antibacterial activity, whereas MS, RM, PR, LG, and LM showed a slightly lower antibacterial activity, and PC, PM, and EC showed an even lower antibacterial activity (Table 2).

Optical density of bacterial cultures incubated with different essential oils at two different concentrations

Our results also showed that TM, MS, RM, PR, and LG had high antibacterial activity at all concentrations, and EOs except for TT and BG showed a concentration-dependent increase in their antibacterial activities (Figure 2). That is, as their concentration increased, the antibacterial activity of MS, RM, PR, LV, LM, PM, and EC also increased.

Figure 2.

OD differences among bacterial cultures in the presence of two different EO concentrations.

Bars represent the differences in OD between cultures grown with the same EO at two different concentrations. The greater the difference in OD values, the higher the concentration-dependent antibacterial activity. EO, essential oil; TM, thyme white; PR, palmarosa; RM, rosemary verbenone; MS, melisa true; LG, lemongrass; RW, rosewood; LV, lavender Bulgarian; LM, Lemon; CP, cypress; CW, cedarwood atias; TT, tea tree; BG, bergamot; OR, orange sweet; SW, sandalwood; PC, pachuli; PM, peppermint premium; EC, eucalyptus bluegum; OD, optical density.

3. Evaluation of antibacterial activity of EOs according to the extraction site

The TM, MS, RM, PR, and LG extracts used here were extracted from leaves; they all showed a high antibacterial activity against E. coli. Therefore, EO antibacterial activity might be related to the extraction site. As previously mentioned, those EOs that had an antibacterial activity similar to that of the antibiotic ampicillin, i.e., TM, PR, and RM, were leaf extracts.

Discussion

In this study, we compared the antibacterial activity of 17 EOs with that of ampicillin against E. coli. Our results showed that TM, PR, and RM had the same antibacterial activity as ampicillin. Moreover, TM had the same antibacterial activity as ampicillin at both 0.5% and 1%. The antibacterial activity of TM may be high due to the presence of carvacrol, γ-terpinene (Alsaraf et al., 2020), and thymol (Kwon et al., 2018) components. It has previously been shown that TM extracted from native plants from Oman has a high content of carvacrol, a compound that bears a higher antibacterial activity than ampicillin (Alsaraf et al., 2020). Our results are in accordance with those from many previous studies showing that TM exhibits an excellent antibacterial activity (Beksac et al., 2021; Sim et al., 2019). Here, it was shown that this activity was not concentration-dependent.

TM also improves atopic dermatitis by inhibiting the activity of Staphylococcus aureus (Alsaraf et al., 2020; Kwon et al., 2018), which may be useful for future antibacterial evaluation of this EO against other bacteria associated with atopic dermatitis.

When the antibacterial activity of EOs was separately evaluated at different concentrations, here and in previous studies, it was observed that TM, PR (da Rocha Neto et al., 2019; Oliveira Ribero et al., 2020), RM (Jawad et al., 2018), LG (Yoon & Park, 2018), and MS (Abdellatif et al., 2021) had high antibacterial activity, that LV and RW had a concentration-dependent antibacterial activity, and that TT did not have any antibacterial activity (Yi & Bu, 2017).

Previous studies have shown that EOs have high antibacterial activity (Puškárová et al., 2017), but high concentrations of EOs do not necessarily have a high antibacterial activity. The concentrations of EO exhibiting an adequate antibacterial activity in some bacteria varies, and it is important to determine the appropriate concentrations and conditions for skin and clinical applications. Here, it was determined that EOs extracted from leaves had very high antibacterial activity PR, RM, MS and LG (Bajalan et al., 2017; Nguyen et al., 2018; Wang et al., 2019), including TM (Gonzalez et al., 2021; Mehrabi et al., 2021), PR, RM (Abozahra et al., 2020), MS (Keskin & Guvensen 2018), and LG (Ilango et al., 2019; Gao et al., 2020). Thought the above five types of EOs extracted from the leaves showed the highest antibacterial activity, LV (Park & Kang, 2020) and LM (Lee et al., 2020a), which were extracted from flowers and fruit skins, also showed high antibacterial activity in a concentration-dependent manner. This is because the composition of essential oils and the mechanism of antibacterial action against bacteria differ depending on the extraction site and botanical family (Kim, 2019). Future research is needed to confirm the effects of EO blending and the appropriate application method for each EO according to an individual's condition, target infection site, and bacterial species.

Conclusions

In this study, the antibacterial activity against E. coli was evaluated for 17 different EOs. First, it was determined that an antibacterial activity similar to that of ampicillin could be found in TM, PR, and RM. Second, among all EOs, TM showed the best antibacterial activity at a concentration of 0.5% (v/v), and TM, PR, and RM showed high antibacterial activity at a 1% (v/v) concentration. Third, EOs with the highest antibacterial activities were mainly extracted from leaves (Lee & Kim, 2021; Kim et al., 2019; Lee et al., 2020c). These results indicate that several EOs have an excellent antibacterial activity.

However, while the antibacterial activity of 17 different EOs was tested, only one antibiotic was used for comparison. Comparisons with a larger group of antibiotics would be a useful complement for the results presented here. Additional studies on effective antibacterial EO concentrations, expressed as MIC, against more than one type of bacterium and resistant strains might also be a good complement for this study.

Despite this, it is meaningful that the antibacterial activity of a number of EOs was evaluated against E. coli, an indicator bacterium, and that the comparison between such antibacterial activity with that of an antibiotic had not been addressed in previous studies. Through research on the antibacterial activities and various properties of EOs, the possibility of useful applications in the field of customized cosmetics preparation and cosmetics manufacturing has been confirmed. Our results may be used as basic data for further research on EOs.

Notes

Author's contribution

JMP and YSY designed, performed experiments, and wrote the manuscript. YKL did the experimental design. JKK has contributed greatly to the review and writing of the manuscript.

Author details

Jung Min Park (Graduate student), Department of Public Health Science, Dankook University, 119, Dandae-ro, Dongnam-gu, Cheonan-si, Chungnam 31116, Korea; Young Sam Yuk (Lecturer), Department of Clinical Medical Science, Dankook University, 119, Dandae-ro, Dongnamgu, Cheonan-si, Chungnam 31116, Korea; Young Ki Lee (Professor) and Jae Kyung Kim (Professor), Department of Biomedical Laboratory Science, Dankook University, 119, Dandae-ro, Dongnam-gu, Cheonan-si, Chungnam 31116, Korea.

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Article information Continued

Figure 1.

Antibacterial activity of different essential oils and ampicillin.

The bars show the antibacterial activity of 17 EOs and ampicillin expressed as the optical density of bacterial cultures. Among the EOs, only TM, PR, and RM showed the same antibacterial activity as ampicillin. The horizontal line marks the results obtained with ampicillin, as a reference. EO, essential oil; TM, thyme white; PR, palmarosa; RM, rosemary verbenone; MS, melisa true; LG, lemongrass; RW, rosewood; LV, lavender Bulgarian; LM, Lemon; CP, Cypress; CW, cedarwood atias; TT, tea tree; BG, bergamot; OR, orange sweet; SW, sandalwood; PC, pachuli; PM, peppermint premium; EC, eucalyptus bluegum.

Figure 2.

OD differences among bacterial cultures in the presence of two different EO concentrations.

Bars represent the differences in OD between cultures grown with the same EO at two different concentrations. The greater the difference in OD values, the higher the concentration-dependent antibacterial activity. EO, essential oil; TM, thyme white; PR, palmarosa; RM, rosemary verbenone; MS, melisa true; LG, lemongrass; RW, rosewood; LV, lavender Bulgarian; LM, Lemon; CP, cypress; CW, cedarwood atias; TT, tea tree; BG, bergamot; OR, orange sweet; SW, sandalwood; PC, pachuli; PM, peppermint premium; EC, eucalyptus bluegum; OD, optical density.

Table 1.

Essential oils used in this study, and tissue/organ extraction sites

Extraction organ/tissue Oil’s name Botanical name Origin Abbreviation
Blossoms Lavender Bulgarian Lavandula vera Bulgaria LV
Leaf Cypress Cupressus sempervirens Austria CP
Eucalyptus bluegum Eucalyptus globulus China EC
Lemongrass Cymbopogon schoenanthus Indonesia LG
Melisa true Melissa officinalis Italy MS
Pachuli Pogostemon cablin Indonesia PC
Palmarosa Cymbopogon martinii India PR
Peppermint premium Mentha piperita India PM
Rosemary Verbenone Rosmarinus officinalis France RM
Tea tree Melaleuca alternifolia Australia TT
Thyme white Thymus vulgaris Spain TM
Peel Bergamot Citrus bergamia Italy BG
Lemon Citrus lemon Sicily LM
Orange sweet Citrus sinensis Brazil OR
Wood Cedarwood atias Cedrus atlantica Nepal CW
Rosewood Aniba rosaeodora Brazil RW
Sandalwood Santalum album India SW

Table 2.

Optical density of bacterial cultures incubated with different essential oils at two different concentrations

No. Oil’s name 0.5% EO
1% EO
Optical density (OD600)* Standard deviation Optical density (OD600) Standard deviation
Ampicillin 0.09
1 Thyme white (TM) 0.11 0.00 0.10 0.00
2 Palmarosa (PR) 0.19 0.13 0.10 0.00
3 Rosemary verbenone (RM) 0.22 0.04 0.10 0.00
4 Melisa true (MS) 0.21 0.03 0.12 0.01
5 Lemongrass (LG) 0.14 0.00 0.13 0.01
6 Rosewood (RW) 0.50 0.02 0.15 0.08
7 Lavender Bulgarian (LV) 0.40 0.01 0.20 0.09
8 Lemon (LM) 0.27 0.06 0.22 0.00
9 Cypress (CP) 0.40 0.03 0.33 0.00
10 Cederwood atias (CW) 0.34 0.00 0.33 0.00
11 Tea tree (TT) 0.37 0.01 0.37 0.01
12 Bergamot (BG) 0.38 0.01 0.38 0.01
13 Orange sweet (OR) 0.40 0.01 0.39 0.01
14 Sandalwood (SW) 0.40 0.01 0.39 0.01
15 Pachuli (PC) 0.46 0.01 0.43 0.01
16 Peppermint premium (PM) 0.61 0.06 0.45 0.03
17 Eucalyptus bluegum (EC) 0.70 0.19 0.55 0.04
*

The OD values for cultures incubated with TM, PR, and RM, depicted in red, are similar to those for ampicillin. As the EO concentration increased, the OD values of cultures decreased.

OD, optical density; EO, essential oil.