Chemical composition and in vitro antibacterial effects of vapours of essential oils from plants recommended by the European Medicines Agency against respiratory infections

Julien Antih

Doctoral thesis

Abstract:

Often used as a synonym for pneumonia, lower respiratory tract infections remain among the leading causes of morbidity and mortality worldwide. Antibiotic inhalation therapy, a method involving the precise delivery of aerosolized particles directly into the lungs, offers distinct advantages over systemic antibiotics. However, challenges in executing effective nebulization techniques, coupled with the lack of robust clinical data, have constrained its broader application, especially in cases of lower respiratory tract infections in children under five. In this context, the volatile constituents of essential oils (EOs) appear as a promising alternative in the development of novel inhaled antibiotic therapy. The purpose of this study was first to determine the in vitro antibacterial activity of five EOs recommended by the European Medicines Agency (EMA) against cough and common cold. The EOs obtained from three commercial samples of Eucalyptus globulus, Foeniculum vulgare, Mentha x piperita, Pimpinella anisum, and Thymus vulgaris were assessed using broth microdilution volatilization (BMV) method against three standard bacterial strains associated with lower tract respiratory infections, including Staphylococcus aureus, Streptococcus pyogenes and Haemophilus influenzae. With the aim to optimize a protocol for characterizing EO vapours, we initially determined the chemical profile of the most active EO (T. vulgaris) using dual column/dual detector gas chromatography system. Subsequently, a thorough characterization of the vapour composition was conducted through headspace analyses above a liquid matrix (LM) of broth and T. vulgaris EO. This was achieved by employing both Solid Phase MicroExtraction (HS-SPME) and Gas Tight Syringe (HS-GTS) sampling techniques. Finally, in our efforts to enhance the antimicrobial activity of T. vulgaris EO vapour, we profiled its composition using a solid matrix (SM) composed of cellulose. The results showed that, out of these 5 EOs, only T. vulgaris possessed a significant activity against the studied respiratory pathogens wvolatile compoundsith MICs comprised in between to 512 and 1024 µg/mL in both liquid and vapour phase. Furthermore, H. influenzae displayed the highest susceptibility to thyme EO, with a MIC of 128 µg/mL. In addition, GC-MS analysis results showed a different distribution of volatile compounds in the headspace. Notably, thymol prevalence was surprisingly low with peak percentage area inferior to 5.27 % (HS-SPME) and 0.60 % (HS-GTS) in the headspace above the LM, as opposed to the EOs (max. 48.65%). Furthermore, oxygenated monoterpenoids were significantly more prevalent in the headspace above the SM – e.g. max. thymol MICs = 43.76 % with HS-SPME and 4.87 % with HS-GTS – than above the LM, whereas hydrocarbon monoterpenes experienced a decrease in abundance. Multiple factors could explain these results such as parameters related to the use of a fibre coating assembly, but also the mechanisms governing the release of volatile compounds into the headspace and their interactions with chosen matrices as well as the degradation reactions occurring in between the volatile substances. Overall, this study holds great promise in providing invaluable insights that could lead to both innovative patents and advancements in pharmaceutical product development. Looking ahead, achieving control over the stability and release of EO volatile substances in vapour presents a formidable challenge in designing a potential antimicrobial vapour inhalation device. However, further investigations, including cytotoxicity studies and clinical aromatherapy trials, are necessary to evaluate the efficacy, tolerability, and safety of EO vapour in treating respiratory diseases.

Keywords

antimicrobial activity, respiratory infections, essential oil, Thymus vulgaris, volatile compounds, headspace analysis

Language used: English

Date on which the thesis was submitted / produced: 11. 1. 2024

Thesis defence

• Supervisor: prof. Ing. Ladislav Kokoška, Ph.D.
• Reader: Pavel Klouček, Ph.D., doc. Ing., Pavlína Kyjaková, externi, Sabrina Boutefnouchet, externi

Czech University of Life Sciences Prague

Faculty of Tropical AgriSciences

Doctoral programme / field:
Agricultural Specialization / Tropical Agrobiology and Bioresource Management