A Comparative Study of Headspace and Hydro-Distillation Techniques for Volatile Compounds in Thyme Species across Drought Stress and Phenological Stages

Document Type : Research Paper

Authors

1 Department of Plant Breeding and Biotechnology, Shahrekord University, Iran

2 Department of Medical Laboratory Science, College of Science, Knowledge University, Kirkuk Road, Erbil 44001, Iraq

Abstract

Variations in essential oils and phenolic compounds of T. daenensis, T. armeniacus, and T. vulgaris were investigated at two phenological stages: 50 and 100% flowering. The volatile components of these species extracted using headspace and hydrodistillation techniques were analyzed via gas chromatography-mass spectrometry (GC-MS) under two water stress conditions (90 and 50% field capacity). GC-MS analysis identified the major constituents, including thymol (5.175–51.53%), carvacrol (1.77–24.52%), γ-terpinene (3.95–23.66%), and p-cymene (3.35–24.50%), in the three species using the hydrodistillation method. Maximum thymol content of 51.53% and 43.20% was recorded in T. daenensis during 100% flowering under 90 and 50% FC conditions, respectively. A decline in thymol content in T. daenensis and T. vulgaris was recorded as a decrease from 90 to 50% FC, while T. armeniacus showed an increase. In the headspace method, different compounds were identified as the primary components. Notably, thymol (7.68–54.34%) was identified as the dominant compound in T. daenensis. Drought stress significantly increased thymol composition at the 50% flowering stage but reduced it at the 100% flowering stage. Hierarchical clustering and principal component analysis revealed three distinct groups, separating the compounds identified by the headspace and hydrodistillation methods. The findings suggest that hydrodistillation is more reliable for determining the true concentrations of compounds.

Keywords

Main Subjects


  1. Aksit H., Bayar Y., Simsek S, Ulutas Y., Chemical Composition and Antifungal Activities of the Essential Oils of Thymus Species (Thymus pectinatus, Thymus convolutus, Thymus vulgaris) Against Plant Pathogens. Journal of Essential Oil-Bearing Plants. 2022;25(1):200–207.
  2. Beicu R., Alexa E., Obiștioiu D., Cocan I., Imbrea F., Pop G., Circioban D., Moisa C., Lupitu A., Copolovici L., Copolovici D.M. Antimicrobial Potential and Phytochemical Profile of Wild and Cultivated Populations of Thyme (Thymus sp.) Growing in Western Romania. Plants. 2021;10(9):2-21.
  3. Semeniuc C.A., Socaciu M.I., Socaci SA., Mureșan V., Fogarasi M., Rotar A.M. Chemometric comparison and classification of some essential oils extracted from plants belonging to Apiaceae and Lamiaceae families based on their chemical composition and biological activities. Molecules. 2018;23(9): 1-13.
  4. Askary M., Behdani M.A., Parsa S., Mahmoodi S., Jamialahmadi M. Water stress and manure application affect the quantity and quality of essential oil of Thymus daenensis and Thymus vulgaris. Industrial Crops and Products. 2018;336–344.
  5. Micucci M., Protti M., Aldini R., Frosini M., Corazza I., Marzetti C., Mattioli L.B., Tocci G., Chiarini A., Mercolini L., Budriesi R. Thymus vulgaris L. essential oil solid formulation: Chemical profile and spasmolytic and antimicrobial effects. Biomolecules. 2020;10(6):860.
  6. Gholami-Ahangaran M., Ahmadi-Dastgerdi A., Karimi-Dehkordi M. Thymol and carvacrol; as antibiotic alternative in green healthy poultry production. Plant Biotechnology Persa. 2020;2(1):22-25.
  7. Stefanakis M.K., Papaioannou C., Lianopoulou V., Philotheou-Panou E., Giannakoula A.E., Lazari D.M. Seasonal variation of aromatic plants under cultivation conditions. Plants. 2022;11(16):1-21.
  8. Soleimani M., Daryasari A.P., Ghorbani A., Mortaz Hejri O., Mazaheri R. Analysis of the Volatile Compounds in Thymus vulgaris L. Using Improved HS-SPME-GC-MS and Comparison with Conventional Methods. Taylor & Francis. 2015;17(6):1233–1240.
  9. Azar P.A., Porgham-Daryasari A., Saber-Tehrani M., Soleimani M. Analysis of the Volatile Compounds in Nepeta crispa Willd. Using Improved HS-SPME–GC–MS and Comparison with Conventional Methods. Acta Chromatographica. 2012.24(1):75-84
  10. Silva L.V., Nelson D.L., Drummond M.F.B., Dufosse L., Gloria M.B.A. Comparison of hydrodistillation methods for the deodorization of turmeric. Food Research International. 2005;38:1087-1096.
  11. Baser KH.C., Buchbauer G. Handbook of essential oils: science, technology, and applications. CRC Press. 2009.
  12. Snow NH. Head-space analysis in modern gas chromatography. Trends in Analytical Chemistry. 2002;21:608-617.
  13. Wilson L.A., Senech, N.P., Widrlechner M.P. Headspace Analysis of the Volatile Oils of Agsstachet. Journal of Agricultural and Food Chemistry. 1992;40:1362-1366.
  14. Friščić M., Maleš Ž., Maleš I., Duka I., Radonić A., Mitić B., Hruševar D., Jurić S., Jerković I. Gas Chromatography–Mass Spectrometry Analysis of Volatile Organic Compounds from Three Endemic Iris Taxa: Headspace Solid-Phase Microextraction vs. Hydrodistillation. Molecules. 2024;29(17):4107.
  15. Nezhadali A., Akbarpour M., Shirvan B.Z., Mousavi M. Comparison of volatile organic compounds of Thymus vulgaris using hydrodistillation and headspace solid phase microextraction gas chromatography mass spectrometry. Journal of the Chinese Chemical Society. 2010;57(1):40-43.
  16. Karami-Osboo R., Miri R., Asadollahi M., Jassbi A.R. Comparison between head-space SPME and hydrodistillation-GC-MS of the volatiles of Thymus daenensis. Journal of Essential Oil Bearing Plants. 2015;18(4):925-930.
  17. Nikolić B., Matović M., Mladenović K., Todosijević M., Stanković J., Đorđević I., Marin PD., Tešević, V. Volatiles of Thymus serpyllum obtained by three different methods. Natural Product Communications. 2019;14(6):1-3.
  18. Abbasi Sh., Houshmand S., Ahmadi T. Volatile compositions and glandular trichomes of Zataria multiflora in different phenological stages under normal and drought stress conditions. BMC Plant Biology. 2024;24:483.
  19. NaghdiBadi H., Yazdani S., Mohammad Ali S.M., Nazari F. Effects of spacing and harvesting time on herbage yield and quality/quantity of oil in thyme, Thymus vulgaris L. Industrial Crops and Products. 2004;19:231-236.
  20. Yousefzadeh K., Houshmand S., Shiran B., Mousavi-Fard S., Zeinali H., Nikoloudakis N., Gheisari M.M., Fanourakis D. Joint Effects of Developmental Stage and Water Deficit on Essential Oil Traits (Content, Yield, Composition) and Related Gene Expression: A Case Study in Two Thymus Species. Agronomy. 2022;12:1-19.
  21. Nickavar B., Mojab F., Dolat-Abadi R. Analysis of the essential oils of two Thymus species from Iran. Food Chemistry. 2005;90:609-611.
  22. Omidbaigi R., Kazemi Sh., Daneshfar E. Harvest Time Effecting on the Essential Oil Content and Compositions of Thymus vulgaris. Journal of Essential Oil Bearing Plants. 2008;11(2):162-167.
  23. Nejad-Ebrahimi S., Hadian J., Mirjalili M.H., Sonboli A., Yousefzadi M. Essential oil composition and antibacterial activity of Thymus caramanicus at different phenological stages. Food Chemistry. 2008;110:927-931.
  24. Wang W.X., Vinocur, B., Shoseyov, O., Altman A. Biotechnology of plant osmotic stress tolerance: physiological and molecular considerations. Acta Horticulturae. 2001;560:285-293.
  25. Tatrai Z.A., Sanoubar R., Pluhar Z., Mancarella S., Orsini F., Gianquinto, G. Morphological and Physiological Plant Responses to Drought Stress in Thymus citriodorus. International Journal of Agronomy. 2016;1-8.
  26. Alavi-Samani S.M., Pirbalouti A.G., Kachouei M.A., Hamdi B. The influence of reduced irrigation on herbage, essential oil yield and quality of Thymus vulgaris and Thymus daenensis. J. of Herb. Drug. 2013;4:109-113.
  27. British Pharmacopoeia. HMSO London, PP .2, A137-A138. 1988.
  28. Adamz R.P. Identification of Essential Oil Components by Gas Chromatography/Mass Spectrometry, Allured Publishing Corporation, Carol Stream. 2007.
  29. Weisany W., Raei Y., Pertot I. Changes in the essential oil yield and composition of dill (Anethum graveolens L.) as response to arbuscular mycorrhiza colonization and cropping system. Industrial Crops and Products. 2015;77:295-306.
  30. Bahreininejad B., Razmjoo J., Mirza M. Influence of water stress on morpho-physiological and phytochemical traits in Thymus daenensis. International Journal of Plant Production. 2013;7(1):151-166.
  31. Tohidi B., Rahimmalek M., Arzani A. Essential oil composition, total phenolic, flavonoid contents, and antioxidant activity of Thymus species collected from different regions of Iran. Food Chemistry. 2017;220:153-161.
  32. Askary M., Talebi S.M., Amini F., Dousti B., Bangan A. Effect of NaCl and iron oxide nanoparticles on Mentha piperita essential oil composition. Environmental and Experimental Biology. 2017;14:27-32.
  33. Kollner T.G., Held M., Lenk C., Hiltpold I., Turlings T.C.J., Gershenzon, J., Degenhardt J.A. maize (E)-β-caryophyllene synthase implicated in indirect defense responses against herbivores is not expressed in most American Maize varieties. Plant Cell. 2008;20:482-494.
  34. Aziz E.E., Hendawy, S.F., Ezz El-Din A.A., Omer E.A. Effect of soil type and irrigation intervals on plant growth, essential oil yield and constituents of Thymus vulgaris plant. American-Eurasian Journal of Agricultural & Environmental Science. 2008;4:443-450.
  35. Ghasemi Pirbalouti A., Rahmani Samani M., Hashemi M., Zeinali H. Salicylic acid affects growth, essential oil and chemical compositions of thyme (Thymus daenensis Celak.) under reduced irrigation. Plant Growth Regulation. 2014;72:289-301.
  36. Stokl J., Brodmann J., Dafni A., Ayasse M., Hansson B.S. Smells like aphids: orchid flowers mimic aphid alarm pheromones to attract hoverflies for pollination. Proceedings of the Royal Society B. 2011;278:1216-1222.
  37. Poulose A.J., Croteau R. Biosynthesis of aromatic monoterpenes: conversion of γ-terpinene to p-cymene and thymol in Thymus vulgaris L. Archives of Biochemistry and Biophysics. 1978;187(2):307-314.
  38. Wesolowska A., Grzeszczuk M., Jadczak D. Comparison of the chemical composition of essential oils isolated by water-steam distillation and hydrodistillation from garden thyme (Thymus vulgaris L.). Journal of Essential Oil Bearing Plants. 2016;19(4):832-842.
  39. Jordan M.J., Martinez R.M., Goodner K.L., Baldwin E.A., Sotomayor A. Seasonal variation of Thymus hyemalis Lange and Spanish Thymus vulgaris L. essential oils compositions. Industrial Crops and Products. 2006;24:253-263.
  40. Sarajuoghi M., Abbaszadeh B., Ardakani M.R. Investigation morphological and physiological response of Thymus vulgaris L. to drought stress. Journal of Biodiversity and Environmental Sciences. 2014;5:486-492.
  41. McGimpsey J.A., Douglas M.H., Van Klink JW., Beauregard DA., Perry NB. Seasonal variation in essential oil yield and composition from naturalized Thymus vulgaris L. in New Zealand. Flavour and Fragrance Journal. 1994;9:347-352.
  42. Mohammadi H., Ghorbanpour M., Brestic M. Exogenous putrescine changes redox regulations and essential oil constituents in field-grown Thymus vulgaris L. under well-watered and drought stress conditions. Industrial. Crops and Products. 2018;22:119-132.
  43. Tohidi B., Rahimmalek M., Arzani A., Trindade H., Sequencing and variation of terpene synthase gene (TPS2) as the major gene in biosynthesis of thymol in different Thymus species. Phytochemistry 2020;169: 1–9.
  44. Abbasi Sh., Houshmand S., Ghorbani S. Gene expression and metabolite analyses of Thymus daenensis and T. vulgaris in the Trichome and leaf at two phenological stages. Biocatalysis and Agricultural Biotechnology. 2024;1-57
  45. Legault J., Pichette A. Potentiating effect of β-caryophyllene on anticancer activity of α‐humulene, isocaryophyllene and paclitaxel. Journal of Pharmacy and Pharmacology. 2007;59:1643-1647.
  46. Burt S. Essential oils: Their antibacterial properties and potential applications in foods-a reviewInternational Journal of Food Microbiology. 2004;94:223-253.
  47. Martinez-Nataren D.A., Parra-Tabla V., Dzib G., Calvo-Irabien L.M. Morphology and density of glandular trichomes in populations of Mexican oregano (Lippia graveolens H.B.K. Verbenaceae), and the relationship between trichome density and climate. Journal of Torrey Botan Society. 2011;138(2):134-144.
  48. Tozin L.R.S., Marques M.O.M., Rodrigues T.M. Glandular trichome density and essential oil composition in leaves and inflorescences of Lippia origanoides Kunth (Verbenaceae) in the Brazilian Cerrado. The Annals of the Brazilian Academy of Sciences. 2015;87(2):943-953.
  49. Ahmadi T., Shabani L., Sabzalian M.R. LED light sources improved the essential oil components and antioxidant activity of two genotypes of lemon balm (Melissa ofcinalis L.). Botanical Studies. 2021;62(1): 1-13.
  50. Shahraki SH., Ahmadi T., Jamali B., Rahimi M. The biochemical and growth associated traits of basil (Ocimum basilicum L.) affected by silver nanoparticles and silver. BMC Plant Biology. 2024;24(1):92.
  51. Yarmoohammadi M., Talebi S.M., Nohooji M.G. Intraspecific variations in essential oil and glandular trichomes in Nepeta heliotropifolia. Biodiversitas Journal. 2017; 18:964-970.