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Analysis of the Hydrosol Aroma of Indian Oregano

Ram S. Verma*, Rajendra C. Padalia and Amit Chauhan
CSIR-Central Institute of Medicinal and Aromatic Plants, Research Centre, Pantnagar, P.O.- Nagla, Dairy farm, Udham Singh Nagar, Uttarakhand-263149, India
Corresponding Author : Ram Swaroop Verma
CSIR-Central Institute of Medicinal and Aromatic Plants
Research Centre, Pantnagar
P. O. Nagla, Dairy farm, Udham Singh Nagar
Uttarakhand-263149, India
Tel: 91-9756021222
Received July 28, 2011; Accepted July 30, 2012; Published August 07, 2012
Citation: Verma RS, Padalia RC, Chauhan A (2012) Analysis of the Hydrosol Aroma of Indian Oregano. Med Aromat Plants 1:112. doi: 10.4172/2167-0412.1000112
Copyright: © 2012 Verma RS, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

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Background: Hydrosols or aqueous distillates are herbal medicinal water obtained as a side product during the process of hydro or steam distillation of aromatic plant materials for getting essential oils. Generally, hydrosols are fragrant due to presence of a fraction of hydrophilic components of essential oils. Typically, in commercial production, the essential oils are decanted after completion of distillation process, whereas the hydrosols are discarded as waste or in few cases cohobated back to the distillation still. Methods: In present study, Indian oregano (Origanum vulgare L.) populations (two chemotypes, viz. thymol, and carvacrol) were hydrodistilled and the compositions of resulted main essential oil (primary oil) and essential oil fraction dissolved in hydrosol (secondary oil) have compared using gas chromatography (GC/FID) and gas chromatography mass spectrometry (GC/MS) techniques. Results: The primary oils of both the investigated populations were dominated by hydrocarbons (monoterpene, sesquiterpene, benzene derivatives; 53.4-57.6%), followed by their oxygenated derivatives (38.1-44.9%). However,secondary oils were mainly dominated by oxygenated compounds (92.0-96.2%). Conclusion: Present study concluded that the hydrosols of O. vulgare populations, a byproduct of essential oil industry, can be used as a potential source of phenolic compounds, viz. thymol and carvacrol.

Origanum vulgare L.; Hydrodistillation; Essential oil; Hydrophilic fraction; Composition
The essential oils or volatile oils are complex mixture of terpenoids (generally C10 and C15) and some other classes of components, isolated from aromatic plants by physical means, e.g. steam-distillation, hydro-distillation, hydro-cum-steam distillation, and hydro-diffusion processes. These oils find extensive application in flavour, perfumery, cosmetic, and pharmaceutical industries. The process of distillation of aromatic plant material yields essential oil as a main product (primary oil or prime oil) and distillate water, also known as hydrosol as a byproduct. Typically, in commercial production, the essential oils are skimmed off and the hydrosol is discarded as waste or cohobated back to the source solution. The slightly hydrophilic part of essential oil, passes in to the aqueous phase during process of distillation, gives pleasant aroma to the resulted hydrosol. Typically, the essential oil fraction present in hydrosol is highly aromatic and its composition is often different from the primary essential oil [1,2]. However, the major components are generally the same of those present in oxygenated fraction of corresponding essential oils [3]. Biological and organoleptic properties of the hydrosols make them useful for food and cosmetic industries [2,4-9]. In addition, hydrosols also find application in biological agriculture against mushrooms, mildew, insects, and for fertilization of soils [10].
In Indian subcontinent, Origanum vulgare L. is locally known as ‘Jungali Tulsi’ or ‘Oregano’ or ‘Himalayan marjoram’. The plant is widely used as a very popular spice. Oregano is of great economic value owing to its various traditional and modern applications. It is used as a traditional remedy to treat various ailments such as whooping and convulsive coughs, digestive disorders, and menstrual problems [11,12]. Dried Origanum leaves and essential oils are used by the flavouring industry in various liqueur formulations, tomato sauces, condiments, in baked goods such as pizzas and salad dressings [13]. The genus, Origanum is known for its huge morphological and chemical diversity [14]. The chemotypic and ontogenic variations occurring in the essential oil composition of Indian oregano (Origanum vulgare L.) have been explored [15-18].
Aqueous distillate volatiles of some Indian aromatic plants have been subjected to phytochemical and antimicrobial studies [1,2,19-24]. However, literature survey revealed that there were no such reports available on emerging aromatic crop, Indian oregano; hence a comparison of the composition of primary (main essential oil) and secondary (dissolved fraction of essential oil) volatile oils of two chemotypes of O. vulgare have been conducted in this research.
Materials and Methods
Plant materials
The fresh samples of O. vulgare chemotypes (I & II) were collected from experimental field of Central Institute of Medicinal and Aromatic Plants, Research Centre, Purara, Uttarakhand on 29th June, 2009 when the plants were in flowering stage. Climatologically, the site falls in temperate region (1250 m) of western Himalaya where the monsoon usually breaking in June and continuing up to September.
Isolation of essential oil
Freshly harvested plant materials were hydro-distilled for 3 hrs in a slightly modified all glass Clevenger’s apparatus in which the recycling of distillate was stopped and diverted to a collection flask so that essential oil and hydrosol can be collected simultaneously (like field distillation without cohobation) [2]. The oil collected directly in extraction burette was separated and dehydrated by anhydrous sodium sulphate and designated as ‘primary oil’. The hydrosol collected simultaneously in a separate flask was used for isolation of ‘dissolved’ or ‘secondary oil’.
Isolation of dissolved essential oil
The hydrosol collected in a separate flask was shaken vigorously with hexane (10:1×2) using separatory funnel for 30 minutes to recover the dissolved oil. The mixture was then allowed to settle and the organic layer was separated. The organic layer was then dried over anhydrous sodium sulphate, filtered and the solvent evaporated (35°C) under reduced pressure to get the ‘secondary oil’. The primary and secondary oils were kept in a cool and dark place prior to analysis.
Gas chromatography (GC)
The GC analysis of the oil sample was carried out on a Nucon gas chromatograph model 5765 equipped with FID and DB-5 (30 m×0.32 mm; 0.25 μm film coating) fused silica capillary column. Oven temperature programming was done from 60-230°C at 3°C/min. Hydrogen was the carrier gas at 1.0 ml/min. The injector and detector temperatures were 220°C and 230°C, respectively. The injection volume was 0.02 μl neat (syringe: Hamilton 1.0 μl capacity, Alltech USA) and the split ratio was 1:30.
Gas chromatography-mass spectrometry (GC/MS)
GC/MS analysis of the essential oil sample was carried out on a PerkinElmer AutoSystem XL GC interfaced with a Turbomass Quadrupole Mass Spectrometer fitted with an Equity-5 fused silica capillary column (60 m × 0.32 mm i.d., film thickness 0.25 μm). The oven temperature was programmed from 60-210°C at 3°C/min using helium as the carrier gas at 1.0 mL/min. The injector temperature was 210°C, injection volume 0.1 μl prepared in n-hexane (dilution 10%), split ratio 1:40. MS were taken at 70 eV with a mass scan range of 40- 450 amu and scan rate 1 sec with interscan delay 0.5 sec.
Identification of components
Constituents were identified on the basis of a Retention Index (RI, determined with reference to homologous series of n-alkanes, C8-C30, under identical experimental conditions), co-injection with standards (Aldrich and Fluka) or known essential oil constituents, MS Library search (NIST/EPA/NIH version 2.1 and WILEY registry of MS data 7th edition), by comparing with the MS literature data [25]. The relative amounts of individual components were calculated based on the GC peak area (FID response) without using a correction factor.
Results and discussion
Primary and secondary volatile oils obtained from the hydrodistillation of fresh herbs of O. vulgare chemotypes (I & II) were analysed by GC/FID and GC/MS. Altogether, 55 constituents representing 95.2-98.3% of the total oil compositions were identified (Table 1). Chemotypes ‘I’ was rich in thymol, p-cymene, and γ-terpinene. While the chemotype II contained carvacrol, p-cymene, and γ-terpinene as major constituents. The amount of thymol and carvacrol was found to be very low in primary oil (30.8% and 1.0%, respectively) as compared to secondary oil (83.4% and 5.9%, respectively) of chemotype ‘I’. On the other hand, primary oil possessed higher amounts of γ-terpinene (21.4%), p-cymene (19.0%), myrcene (3.2%), carvacrol methyl ether (3.2%), α-terpinene (2.4%), (E)-caryophyllene (2.1%), (E)-β-ocimene (2.0%), and (Z)-β-ocimene (1.3%) as compared to secondary oil of this chemotype. Furthermore, the primary oil of chemotype ‘II’ contained lesser amount of carvacrol (42.3%) in comparison to secondary oil (94.7%). However, the amounts of γ-terpinene (23.4%), p-cymene (20.9%), myrcene (2.0%), and α-terpinene (2.0%) were noticed to be higher in primary oil than secondary oil of chemotype II.
Further, the class compositions of the primary and secondary volatile oils of O. vulgare chemotypes also showed clear and considerable differences in their nature. The primary oils of O. vulgare chemotypes were dominated by hydrocarbons (chemotype I: 57.6%; chemotype II: 53.4%), followed by oxygenated compounds (chemotype I: 38.1%; chemotype II: 44.9%). However, secondary oils were dominated only by oxygenated compounds (chemotype I: 92.0%; chemotype II: 96.2%). These variations are observed due to relatively higher solubility of oxygenated compounds in water over the solubility of hydrocarbons.
Thus, on the basis of present study, it can be said that the hydrosol of O. vulgare populations should not be discarded as usually done with commercial distillation of aromatic crops. It could be redistilled by introducing cohobation system in field distillation unit to improve the organoleptic property of the primary oil or reused for distillation of fresh herb to minimize the loss of valuable components of the essential oil. Alternatively, hydrosols could also be used for isolation of pure compounds (thymol, and carvacrol) or it can be used as such for disinfection and cosmetic applications.
We are thankful to Director, CSIR-Central Institute of Medicinal and Aromatic Plants, Lucknow, India for continuous support.

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