![INTRODUCTION
Essential oils (EOs) are characterized as volatile secondary metabolites of plants that provide a unique
aroma, flavor, or both.
More than 17,500 species from various angiosperm families, such as Lamiaceae, Rutaceae, Myrtaceae,
Zingiberaceae, and Asteraceae, produce EOs, but only around 300 of these are used commercially[1].
Although EOs often consist of two or three primary components, making up 20–70% of their
composition, they are highly complex mixtures primarily comprising terpenes, terpenoids, and
phenylpropanoids.
Additionally, they may include a variety of other compounds such as fatty acids, oxides, and sulfur
derivatives [2].
Essential oils (EOs) are typically extracted through methods such as hydrodistillation, steam distillation,
dry distillation, or mechanical cold pressing of botanical materials.
On a laboratory scale, the traditional approach utilizes the Clevenger steam distillation apparatus, which
was invented in 1928.](https://image.slidesharecdn.com/zee-diss1-250604190953-477c0492/85/Amylase-production-in-scienceEE-DISS-1-pptx-3-320.jpg)
![INTRODUCTION (CONTD.)
Humans have utilized essential oils for thousands of years, not just as components in perfumes or for
flavoring food, but also in traditional medicine, due to their various biological attributes, including their
antimicrobial effects [6]
These antimicrobial properties are crucial in addressing the growing challenge of drug-resistant
microorganisms.
Lemongrass is grown worldwide, and India holds a dominant position in its production and export.[10]
As drug resistance develops and the availability of effective treatments for infectious diseases declines,
researchers are increasingly interested in medicinal plants as a source of new compounds with strong
antimicrobial properties.
Recent research indicates that essential oils from plants exhibit significant antimicrobial effects against
various foodborne pathogens, suggesting they have promising potential for application in food
preservation.](https://image.slidesharecdn.com/zee-diss1-250604190953-477c0492/85/Amylase-production-in-scienceEE-DISS-1-pptx-4-320.jpg)
![Collection of plant material
and extraction through
standard methods
Estimation of yield and
quality characteristics
Screening of selected
essential oils for their
antibacterial and antifungal
properties.
By agar well diffusion method
Determination of MIC of EOs
Basic methodology involved
Name of the instrument Manufacturing Company
Autoclave NK Sci & Electrical Engineering, India
Centrifuge (R 24) Remi, India
Chlorophyll Fluorometer (JUNIOR PAM-2000) Walz, Effeltrich, Germany
Distillation Unit (Clevenger’s Apparatus) Borosil, India
Electric air-force oven Jindal S.M.I, India
Electronic Balance (AY 220) Shimadzu, Japan
Fourier-transform Infrared (FT-IR) Spectrometer (2020) Perkin Elmer, USA
Fourier-transform Nuclear Magnetic Resonance (FT-NMR)
Spectrometer
Bruker, USA
Gamma Radiation Chamber
(Co-60 Gamma Irradiator)
Bhabha Atomic Research Centre, Mumbai, India
Gas Chromatograph-Mass Spectrometer (GC-MS) Shimadzu QP-2010 Plus, Japan
Gel Permeation Chromatograph (GPC) Hitachi, Japan
Incubator Jindal S.M.I, India
Infrared gas analyser (IRGA) [LI-COR 6400] Lincoln, NE, USA
MALDI- TOF MS, UltrafleXtreme Bruker Daltonics, USA
Cross-Polarization Magic Angle Spinning Carbon-13 Nuclear
Magnetic Resonance(CP-MAS 13
C- NMR) spectroscopy
JEOL-ECX-400, USA
Scanning Electron Microscope (JSM6510 LV) JEOL, Japan
Sprayer Machine Nature Gold, India
UV- VIS Spectrophotometer (UV-1700) Shimadzu, Japan
Water Distillation Unit Borosil, India
Weighing Balance Square, India](https://image.slidesharecdn.com/zee-diss1-250604190953-477c0492/85/Amylase-production-in-scienceEE-DISS-1-pptx-6-320.jpg)
![RESULTS
Fig 4 and Table 4: Chromatogram of Curry leaves essential oil demonstrating abundance of compounds via GC-MS
S.n
o
Name Area (in
%)
1 BICYCLOHEXANE, 4-METHYLENE-1-
(1-METHYLETHYL)-
15.50
2 Mahanimbine 15.10
3 TRANS (. BETA.)-CARYOPHYLLENE 8.47
4 BICYCLO[3.1.1] HEPT-2-ENE, 2,6,6-
TRIMETHYL-
4.36
5 3-Cyclohexen-1-ol, 4-methyl-1-(1-
methylethyl)-, (R)-
4.25](https://image.slidesharecdn.com/zee-diss1-250604190953-477c0492/85/Amylase-production-in-scienceEE-DISS-1-pptx-11-320.jpg)
![REFERENCES
Mérillon, J.-M.; Rivière, C. Natural Antimicrobial Agents; Springer International Publishing AG: Cham, Switzerland, 2018.
2. Stringaro, A.; Colone, M.; Angiolella, L. Antioxidant, antifungal, antibiofilm, and cytotoxic activities of Mentha spp. essential oils. Medicines 2018,
5, 112. [CrossRef] [PubMed]
3. Deryng, J. Nowyaparat do oznaczanieolejków w materialero´slinnym. Acta Pol. Pharm. 1951, 8, 121–136. Molecules 2019, 24, 2130 15 of 21
4. Arora, R.; Singh, B.; Vig, A.P.; Arora, S. Conventional and modified hydrodistillation method for the extraction of glucosinolate hydrolytic products:
A comparative account. SpringerPlus 2016, 5, 479. [CrossRef] [PubMed]
5. Baj, T.; Sieniawska, E.; Kowalski, R.; Wesolowski, M.; Ulewicz-Magulska, B. Effectiveness of the deryng and clevenger-type apparatus in isolation
of various types of components of essential oil from the MutelinapurpureaThell. flowers. Acta Pol. Pharm. 2015, 72, 507–515. [PubMed]
6. Brnawi, W.I.; Hettiarachchy, N.S.; Horax, R.; Kumar-Phillips, G.; Ricke, S. Antimicrobial activity of leaf and bark cinnamon essential oils against
Listeria monocytogenes and Salmonella typhimurium in broth system and on celery. J. Food Process Preserv. 2019, e13888. [CrossRef]
7. Benzaid, C.; Belmadani, A.; Djeribi, R.; Rouabhia, M. The effects of Mentha × piperita essential oil on C. albicans growth, transition, biofilm
formation, and the expression of secreted aspartyl proteinases genes.Antibiotics 2019, 8, 10. [CrossRef]
8. Brun, P.; Bernabè, G.; Filippini, R.; Piovan, A. In vitro antimicrobial activities of commercially available tea tree (Melaleuca alternifolia) essential
oils. Curr.Microbiol. 2019, 76, 108–116. [CrossRef]
9. Shah, G.; Shri, R.; Panchal, V.; Sharma, N.; Singh, B.; Mann, S. Scientific basis for the therapeutic use of Cymbopogoncitratus, stapf (Lemon grass).
J. Adv. Pharm. Technol. Res. 2011, 2, 3–8. [CrossRef]
10. Srivastava, V.; Dubey, S.; Mishra, A. A Review on Lemon Grass: Agricultural and Medicinal Aspect. Int. Res. J. Pharm. 2013, 4, 42–44. [CrossRef]](https://image.slidesharecdn.com/zee-diss1-250604190953-477c0492/85/Amylase-production-in-scienceEE-DISS-1-pptx-13-320.jpg)
Amylase production in scienceEE-DISS (1).pptx
- 1. Extraction, Characterization, and Antimicrobial Activity of Essential Oil from Lemon Grass (Cymbopogon citratus) and Curry Leaf (Murraya koenigii) Presented by Zeeshan Iqbal (Enrolment no. 2300102234) DEPARTMENT OF BIOENGINEERING, FACULTY OF ENGINEERING &INFORMATION TECHNOLOGY, INTEGRAL UNIVERSITY, LUCKNOW Dissertation on Submitted in partial fulfillment for the award of the degree of M.Tech in Biotechnology Under the Supervision of Prof. Iqbal Ahmad (Supervisor) Dr. Khan Bilal M. Ahmad (Co- Supervisor) Chairman, Dept. of Agricultural Microbiology CAE, Faculty of Agricultural
- 2. CONTENTS Introduction Objectives Methodology Results Conclusion References
- 3. INTRODUCTION Essential oils (EOs) are characterized as volatile secondary metabolites of plants that provide a unique aroma, flavor, or both. More than 17,500 species from various angiosperm families, such as Lamiaceae, Rutaceae, Myrtaceae, Zingiberaceae, and Asteraceae, produce EOs, but only around 300 of these are used commercially[1]. Although EOs often consist of two or three primary components, making up 20–70% of their composition, they are highly complex mixtures primarily comprising terpenes, terpenoids, and phenylpropanoids. Additionally, they may include a variety of other compounds such as fatty acids, oxides, and sulfur derivatives [2]. Essential oils (EOs) are typically extracted through methods such as hydrodistillation, steam distillation, dry distillation, or mechanical cold pressing of botanical materials. On a laboratory scale, the traditional approach utilizes the Clevenger steam distillation apparatus, which was invented in 1928.
- 4. INTRODUCTION (CONTD.) Humans have utilized essential oils for thousands of years, not just as components in perfumes or for flavoring food, but also in traditional medicine, due to their various biological attributes, including their antimicrobial effects [6] These antimicrobial properties are crucial in addressing the growing challenge of drug-resistant microorganisms. Lemongrass is grown worldwide, and India holds a dominant position in its production and export.[10] As drug resistance develops and the availability of effective treatments for infectious diseases declines, researchers are increasingly interested in medicinal plants as a source of new compounds with strong antimicrobial properties. Recent research indicates that essential oils from plants exhibit significant antimicrobial effects against various foodborne pathogens, suggesting they have promising potential for application in food preservation.
- 5. OBJECTIVES Based on the problem of drug resistance mentioned previously and the effectiveness of Essential oils in antimicrobial therapy, our study has been designed with the following objectives 1. To find the effectiveness of the essential oil of lemongrass for the treatment of pathogenic organisms 2. To extract lemon grass and Curry Leaf Essential Oil and Characterization using GC-MS. 3. To test the Activity of Lemon Grass and Curry Leaf Essential Oil on Bacteria 4. To test the Activity of Lemon Grass and Curry Leaf Essential Oil on Fungi
- 6. Collection of plant material and extraction through standard methods Estimation of yield and quality characteristics Screening of selected essential oils for their antibacterial and antifungal properties. By agar well diffusion method Determination of MIC of EOs Basic methodology involved Name of the instrument Manufacturing Company Autoclave NK Sci & Electrical Engineering, India Centrifuge (R 24) Remi, India Chlorophyll Fluorometer (JUNIOR PAM-2000) Walz, Effeltrich, Germany Distillation Unit (Clevenger’s Apparatus) Borosil, India Electric air-force oven Jindal S.M.I, India Electronic Balance (AY 220) Shimadzu, Japan Fourier-transform Infrared (FT-IR) Spectrometer (2020) Perkin Elmer, USA Fourier-transform Nuclear Magnetic Resonance (FT-NMR) Spectrometer Bruker, USA Gamma Radiation Chamber (Co-60 Gamma Irradiator) Bhabha Atomic Research Centre, Mumbai, India Gas Chromatograph-Mass Spectrometer (GC-MS) Shimadzu QP-2010 Plus, Japan Gel Permeation Chromatograph (GPC) Hitachi, Japan Incubator Jindal S.M.I, India Infrared gas analyser (IRGA) [LI-COR 6400] Lincoln, NE, USA MALDI- TOF MS, UltrafleXtreme Bruker Daltonics, USA Cross-Polarization Magic Angle Spinning Carbon-13 Nuclear Magnetic Resonance(CP-MAS 13 C- NMR) spectroscopy JEOL-ECX-400, USA Scanning Electron Microscope (JSM6510 LV) JEOL, Japan Sprayer Machine Nature Gold, India UV- VIS Spectrophotometer (UV-1700) Shimadzu, Japan Water Distillation Unit Borosil, India Weighing Balance Square, India
- 7. Bacteria Lemongrass oil Curry Leaves oil Peppermint oil Eucalyptus oil Clove oil Tetracycline30 DMSO Bacilluscereus MTCC 430 24 22 14 16 20 24 Klebsiella pneumoniae ATCC 1705 18 18 - 15 19 20 Pseudomonas aeruginosa (PAO1) 22 18 - 14 15 19 - Staphylococcus aureus MTCC 737 26 23 12 13 16 22 - Escherichia coli ATCC 25922 (positive control) 27 26 16 18 19 26 - Fungus Lemongrass oil Curry Leaves oil Peppermint oil Eucalyptus oil Clove oil Amphotericin DMSO Candida albicans MTCC 3017 24 21 14 14 16 26 - RESULTS Table 1: Antibacterial and antifungal activity exhibited by essential oils against reference strains
- 8. Lemongrass oil Curry Leaves oil Peppermint oil Eucalyptus oil Clove oil 0 5 10 15 20 25 30 B. cereus K. pneumoniae P. aeruginosa S. aureus E.coli C. albicans Essential oils Zone of inhibition RESULTS Fig. 1:Antimicrobial activity of essential oils against reference strains
- 9. MIC in (µl/mL) Reference strains Lemongrass essential oil Curry leaves essential oil Bacillus cereus MTCC 430 0.5 1.0 Klebsiella pneumoniae ATCC 1705 2.0 2 Pseudomonas aeruginosa (PAO1) 2.0 4 Staphylococcus aureus MTCC 737 0.5 2.0 Escherichia coli ATCC 25922 0.5 1 Candida albicans MTCC 3017 1.0 4.0 Bacillus cereus Klebsiella pneumoniae Pseudomonasaeruginosa Staphylococcus aureus Escherichia coli Candida albicans 0 0.5 1 1.5 2 2.5 3 3.5 4 4.5 Lemongrass oil Curry Leaves oil Reference strains MIC (µl/mL) Table 2 & Fig. 2: MIC exhibited by Lemongrass and Curry leaves essential oil against a panel of drug-resistant pathogens RESULTS
- 10. RESULTS S.no Name Area (in %) 1 Citral 45.76 2 2,6-Octadienal, 3,7- dimethyl- 39 3 Isogeranial 3.44 4 Isogeranial 2.48 5 Limonene 1.94 Fig 3 and Table 3: Chromatogram of Lemongrass essential oil demonstrating abundance of compounds via GC-MS
- 11. RESULTS Fig 4 and Table 4: Chromatogram of Curry leaves essential oil demonstrating abundance of compounds via GC-MS S.n o Name Area (in %) 1 BICYCLOHEXANE, 4-METHYLENE-1- (1-METHYLETHYL)- 15.50 2 Mahanimbine 15.10 3 TRANS (. BETA.)-CARYOPHYLLENE 8.47 4 BICYCLO[3.1.1] HEPT-2-ENE, 2,6,6- TRIMETHYL- 4.36 5 3-Cyclohexen-1-ol, 4-methyl-1-(1- methylethyl)-, (R)- 4.25
- 12. CONCLUSIONS & FUTURE SCOPE Essential oils of Lemongrass and Curry leaves exhibit broad–spectrum activity against fungi and bacteria of clinical importance. The essential oil of Lemongrass demonstrated significantly lower MIC as compared to Curry leaves. Citral, 2,6-Octadienal, 3,7-dimethyl-, Isogeranial, and Limonene are the major compounds present in Lemongrass essential oil as revealed by GC-MS analysis. Major compounds present in Curry leaves essential oil are Bicyclo Hexane, 4-Methylene-1-(1-Methylethyl), Mahanimbine, Trans (Beta.)-Caryophyllene, Bicyclo Hept-2-Ene, 2,6,6-Trimethyl-, 3-Cyclohexen-1-Ol, 4- Methyl-1-(1-Methylethyl) as revealed by GC-MS analysis. The overall findings suggest that essential oils of Lemongrass and Curry leaves are a promising candidate as an alternative agent in antimicrobial therapy against drug-resistant pathogens. Synergistic interaction with antibiotic/s may be studied to further exploit their potential in combinational therapy. Further studies on a suitable model can be conducted to evaluate the efficacy of these essential oils in vivo.
- 13. REFERENCES Mérillon, J.-M.; Rivière, C. Natural Antimicrobial Agents; Springer International Publishing AG: Cham, Switzerland, 2018. 2. Stringaro, A.; Colone, M.; Angiolella, L. Antioxidant, antifungal, antibiofilm, and cytotoxic activities of Mentha spp. essential oils. Medicines 2018, 5, 112. [CrossRef] [PubMed] 3. Deryng, J. Nowyaparat do oznaczanieolejków w materialero´slinnym. Acta Pol. Pharm. 1951, 8, 121–136. Molecules 2019, 24, 2130 15 of 21 4. Arora, R.; Singh, B.; Vig, A.P.; Arora, S. Conventional and modified hydrodistillation method for the extraction of glucosinolate hydrolytic products: A comparative account. SpringerPlus 2016, 5, 479. [CrossRef] [PubMed] 5. Baj, T.; Sieniawska, E.; Kowalski, R.; Wesolowski, M.; Ulewicz-Magulska, B. Effectiveness of the deryng and clevenger-type apparatus in isolation of various types of components of essential oil from the MutelinapurpureaThell. flowers. Acta Pol. Pharm. 2015, 72, 507–515. [PubMed] 6. Brnawi, W.I.; Hettiarachchy, N.S.; Horax, R.; Kumar-Phillips, G.; Ricke, S. Antimicrobial activity of leaf and bark cinnamon essential oils against Listeria monocytogenes and Salmonella typhimurium in broth system and on celery. J. Food Process Preserv. 2019, e13888. [CrossRef] 7. Benzaid, C.; Belmadani, A.; Djeribi, R.; Rouabhia, M. The effects of Mentha × piperita essential oil on C. albicans growth, transition, biofilm formation, and the expression of secreted aspartyl proteinases genes.Antibiotics 2019, 8, 10. [CrossRef] 8. Brun, P.; Bernabè, G.; Filippini, R.; Piovan, A. In vitro antimicrobial activities of commercially available tea tree (Melaleuca alternifolia) essential oils. Curr.Microbiol. 2019, 76, 108–116. [CrossRef] 9. Shah, G.; Shri, R.; Panchal, V.; Sharma, N.; Singh, B.; Mann, S. Scientific basis for the therapeutic use of Cymbopogoncitratus, stapf (Lemon grass). J. Adv. Pharm. Technol. Res. 2011, 2, 3–8. [CrossRef] 10. Srivastava, V.; Dubey, S.; Mishra, A. A Review on Lemon Grass: Agricultural and Medicinal Aspect. Int. Res. J. Pharm. 2013, 4, 42–44. [CrossRef]