Volatile Oil Composition From Flowers and Leaves of H.officinalis L. Grown in Esfahan Headsapce Gas Chromatography/Mass Spectrometry

Abstract

Abstract
 
The aim of this study was to analyze the chemical composition of essential oils from flowers and leaves of Hyssopus officinalis L. from Esfahan province, Iran. The volatile compounds were investigated using Headspace Gas Chromatography/Mass Spectrometry in flowering stage. Twenty nine and twenty seven compounds constituting 99.84% and 99.70 of the oil were identified in the essential oil of flowers and leaves of Hyssopus officinalis L. in flowering stage, respectively. Our results demonstrated that in the flowering stage of H. officinalis L. the major constituents grown in Isafahan of essential oil from the flowers and leaves respectively were Beta-pinene (23.38%– and 31.2%), trans-Pinocamphone (19.34% and, 15.5%), Cis-pinocamphone (27.49% and, 28.1%), myrcene (6.01% and, 6.0 %) and Sabinene (5.89% and, 6.4%)
 
Keywords: Hyssopus officinalis L., Volatile compounds, Headspace GC–MS, Beta-pinene, trans-pinocamphone, Cis-pinocamphone, Myrcene , Sabinene.
 

Keywords


                 Research on Crop Ecophysiology                                  Vol.12/1, Issue 1 (2017), Pages: 37 -  43

 

 

 

Original Research

 

                                                                                                 

Volatile Oil Composition From Flowers and Leaves of H.officinalis L. Grown in Esfahan Headsapce Gas Chromatography/Mass Spectrometry

 

Lida Hashemi*1 and Sayed Komeil  Sayedshourbalal2

1-PhD Student of genetic and Breeding, Department of Agronomy and Plant Breeding, Khorasgan (Isfahan) Branch, Islamic Azad University, Isfahan, Iran

2-PhD Student of agronomy, Department of Agronomy and Plant Breeding, Khorasgan (Isfahan) Branch, Islamic Azad University, Isfahan, Iran

 

 

* Corresponding author email: lida.hashemi@gmail.com

 

Received: 23 January 2017                                  Accepted: 2 February 2017

 

Abstract

 

The aim of this study was to analyze the chemical composition of essential oils from flowers and leaves of Hyssopus officinalis L. from Esfahan province, Iran. The volatile compounds were investigated using Headspace Gas Chromatography/Mass Spectrometry in flowering stage. Twenty nine and twenty seven compounds constituting 99.84% and 99.70 of the oil were identified in the essential oil of flowers and leaves of Hyssopus officinalis L. in flowering stage, respectively. Our results demonstrated that in the flowering stage of H. officinalis L. the major constituents grown in Isafahan of essential oil from the flowers and leaves respectively were Beta-pinene (23.38%– and 31.2%), trans-Pinocamphone (19.34% and, 15.5%), Cis-pinocamphone (27.49% and, 28.1%), myrcene (6.01% and, 6.0 %) and Sabinene (5.89% and, 6.4%)

 

Keywords: Hyssopus officinalis L., Volatile compounds, Headspace GC–MS, Beta-pinene, trans-pinocamphone, Cis-pinocamphone, Myrcene , Sabinene.

 

 

Introduction

 

Plants are important sources for the discovery of new products of medicinal compounds for drug development and plants secondary metabolites are unique sources for pharmaceuticals food additives, flavors and others industrial uses either as part of a final product or as a raw material (Zhao et al., 2005). One of the most frequently consumed herbal remedies available today is the hyssop preparations prepared from Hyssopus officinalis (L.) which is gaining increased importance as a minty flavor, condiment and spices in food industries as well (Dragland et al., 2003; Jung et al., 2004; Lugasi et al., 2006; Wesołowska et al., 2010). Hyssop (Hyssopus officinalis L.) belongs to the Lamiaceae family and is a native plant from central and Southern Europe, Western Asia, and North Africa (Mitić and Đorđević, 2000; Lawless, 2002). The plant is a typical xerophyte and is well adapted to drought and low input conditions (Hornok, 1992).  Medicinal herb, hyssop has a long history of medicinal use as viral infections such as colds, coughs, sore throats, bronchitis and asthma. The oil is antimicrobial, mildly spasmolytic and exhibit strong antiviral activity against HIV (Gollapudi et al., 1995). Antibacterial (Michalczyk et al., 2012), anti-fungal (Fraternale et al., 2004) and antioxidant property of hyssop has been attributed to the presence of pinocamphone, iso-pinocamphone and β-pinene. Antiviral activity has probably been attributed to the presence of caffeic acid and tannins (Gollapudi et al., 1995; Grzeszczuk and Jadczak, 2009). Pinocamphone (trans-Pinocamphone) and iso-pinocamphone (cis-pinocamphone) are generally known as the main characteristic components of the oils of Hyssopus officinalis (Mazzant et al., 1998) contributing approximately 36 to 41% of the total extract. Furthermore, the recommended levels of β-pinene cis-pinocamphone and the second isomer of pinocamphone (trans-pinocamphone) are 13.5- 23% , 34.5 – 50% and 5.5 – 17.5 %, respectively (Mazzant et al., 1998). In some studies, iso-pinocamphone, pinocamphone, β-pinene and pinocarvone were reported to be the most abundant components in hyssop oil (Kizil et al., 2008; De Martino et al., 2009; Veres et al, 1997; Shah et al., 1986; Garg et al., 1999). Generally, Essential oils are complex mixtures of volatile, lipophilic and odiferous substances from the secondary metabolism of plants. They are mainly composed of monoterpenes, sesquiterpenes and their oxygenated derivatives.

Salma (2002) identified H. officinalis as a new source of essential oil in Egypt that was characterized by high content of β-pinene (19.60%), pinocamphone (19.20%) and camphor (16.3%). The highest yield of oil production was determined at the flowering stage of growth in July (Salma. 2002). Different compounds have been identified as the main component in hyssop oil by other researchers. Furthermore, the presence of aliphatic fatty acids, such as palmitic acid (15.60%), stearic acid (10.73%), linolenic acid (63.98%), arachidic acid (2.64%) and eicosadienoic acid 0.68% in the Romanian hyssop oil was determined (Benedec et al., 2002). Özer et al. (2005) analyzed the essential oil of Hyssopus officinalis L. subsp. angustifolius (Bieb.). The essential oil of this plant demonstrated the presence of many monoterpenes that were identified by gas chromatography; about thirty-four components were characterized, representing 91.0% of the total components detected. The main components were identified as pinocarvone, pinocamphone, β-pinene, 1,8-cineole and isopinocamphone (Hold and Sirisoma, 2002; Ozer et al., 2005). H. officinalis var. angustifolius (Persian name: “Zoofa”) is grown and cultivated in some parts of Iran. The aerial parts of hyssop are used in Iranian folk medicine for their asthma, bronchitis, ulcers and wounds, carminative, antiseptic and antimicrobial (Zargari 1990; Ghasemi Pirbalouti, 2009).

Kazazi et al. (2007) reported that  the main components of the extracts under different SFE conditions from H. officinalis cultivated in Iran were sabinene (4.2–17.1%), iso-pinocamphene (0.9–16.5%) and pinocamphene (0.7–13.6%). Detailed examination of the SFE of the hyssop oil was undertaken by Kazazi et al. (2007) at various pressures, temperatures, extraction (dynamic and static), times and modifier (methanol) concentrations. Considering the impacts of different factors during the extraction, it was shown that the composition of the extracted oils was significantly influenced by the operating conditions. Major components of the extracts under different SFE conditions were sabinene (4.2 to 17.1%, w/w), iso-pinocamphene (0.9 to 16.5%) and pinocamphene (0.7 to 13.6%). On the whole, Volatile oil composition varies in dependence on variety, growth stage on the date of collection, climatic conditions and also is affected by extraction and isolation method, and agrotechnical factors (Benhammou et al., 2008; Ghalem and Mohamed, 2009; Xu et al., 2011). Additionally, many plants have various chemotypes that differ in their both quantitative and qualitative diversity in the composition of essential oils obtained (Varga et al., 1998). This work was concluted the chemical composition of essential oils from flowers and leaves of H. officinalis L. at flowering stage using Headspace Gas Chromatography/Mass Spectrometry.

 

Materials and methods

 

Plant Material

 

H. officinalis seeds obtained from the Pakan Seed Company, Isfahan, Iran. Were grown on 26th March, 2017, in plastic greenhouse conditions. Six seeds were sown in each plastic pot filled with clay-loamy soil and after six weeks were thinned to two healthy seedlings per pot. On May 14, 2017, the pots transferred to the Research Field of Islamic Azad University, Branch, Isfahan (Khorasgan), Iran. On July 8, 2017, healthy leaves and flowers were collected from 10 different plants for analysis of essential oils.

 

Identification of the oil components

 

Headspace GC/MS technique was used to analyze volatile compounds. A fresh sample was placed in a closed sampling vessel, heated using a known temperature profile, and the vapor in the vessel was sampled for analysis.

 

GC–MS Analysis

 

An Agilent model 7890 GC interfaced to a 5975C mass selective detector was used for mass spectral identification of the components of the oils. HP-5MS capillary columns (60 m × 0.25 mm × 0.25 μm film thick-ness) were used for GC. The oven temperature was maintained at 40°C for 3 min then programmed to 290°C at 5° C min−1 and remind for 5 min. The carrier gas was helium, at a flow rate of 1.3 mL min−1, and the injection volume was 250 μL. In mass spectrometry electron-impact ionization was performed at electron energy of 70 eV. MS interface temperature was 280°C, and the mode was EI. Detector voltage, mass range scan speed and interval 0.01 min (20 Hz) were, 1.66 Kv, 30 to 550 u, 2.86 scans/s, respectively.

 

 Identification of components

 

The constituents of the volatile oils were also identified by comparing their GC retention indices. A mixture of aliphatic hydrocarbons (C8–C24) in hexane (Sigma–Aldrich, St. Louis, USA) was injected under the above-mentioned temperature programmed to calculate the retention indices. Compound identification was based on the comparison of retention indices using a MS library. The NIST and Wiley spectrometer data bank was used to determine the percentage composition of the compounds.

 

Table1. Chemical composition (%) of essential oil in flowers and leaves.of Hyssopus officinalis L.

 

Compound*

Molecular

formula

Class

Ki Cal

%flower

% leaf

1

alpha-thujene

C10H16

MH

930

1.25

1.2

2

1R-alpha-pinene

C10H16

MH

939

1.46

2.0

3

camphene

C10H16

MH

958

0.37

0.5

4

Sabinene

C10H16

MH

981

5.89

6.4

5

beta-pinene

C10H16

MH

989

23.38

31.2

6

myrcene

C10H16

MH

994

6.01

6.0

7

α-Phellandrene

C10H16

MH

1016

0.11

0.1

8

alpha-terpinene

C10H16

MH

1026

0.98

0.3

9

limonene

C10H16

MH

1040

2.04

2.0

10

beta-phellandrene

C10H16

MH

1043

1.90

2.2

11

β-Ocimene

C10H16

MH

1053

4.74

1.7

12

gamma-terpinene

C10H16

MH

1068

1.38

0.4

13

cis-beta-terpineol

C10H18O

MH

1089

0.26

0.1

14

alpha-terpinolene

C10H16

MH

1095

0.29

0.1

15

Linalool

C10H18O

MH

1116

0.28

0.2

16

β-Thujone

C10H16O

OM

1120

0.24

0.1

17

E,E-alloocimene

C10H16

MH

1137

0.34

0.1

18

Myrtenyl methyl ether

C11H18O

MH

1166

0.33

0.5

19

trans-Pinocamphone

C10H16O

OM

1181

19.34

15.5

20

isopinocamphone

C10H16O

OM

1196

27.49

28.1

21

terpinen-4-ol

C10H18O

MH

1202

0.41

0.2

22

Myrtenol

C15H24

SH

1221

0.26

0.2

23

2,5-Dimethyl-3-methylene-hepta-1,5-diene

C10H16

MH

1342

0.24

0.1

24

α-Gurjunene

C15H24

SH

1424

0.05

-

25

β-Gurjunene

C15H24

SH

1435

0.10

0.2

26

beta-caryophyllene

C15H24

SH

1442

0.09

0.1

27

Germacrene D

C15H24

SH

1450

0.12

-

28

α-Humulene

C15H24

SH

1518

0.14

0.1

29

Oleic Acid

C18H34O2

Fatty Acyls

2077

0.33

0.2

 

Total percentage

 

 

 

99.84

99.70

*The compounds have been arranged according to retention indices relative to (C8-C24) hexane on an HP-5MS capillary column. Ki: Kovatz retention indices given in the literature, MH: Monoterpene hydrocarbons, OM: Oxygenated monoterpene, SH; Sesquiterpene hydrocarbons

 

 

 

Results and discussion

 

The constituents of the obtained essential oils of Hyssopus officinalis L. are presented in Table 1. The Headspace GC/MS analysis method revealed several monoterpenoid hydrocarbons (MH), oxygenated monoterpenes (OM), sesquiterpenoid hydrocarbons (SH). Twenty nine constituents were identified in the essential oil of flowers. The major components were beta-pinene (23.38%), trans-Pinocamphone (19.34%), Cis-pinocamphone (27.49%), myrcene (6.01%) and Sabinene (5.89%). Twenty seven constituents were identified in the essential oil of the leaves and the major components were the same as of the flowers (31.2; 15.5; 28.1; 6.0 and 6.4%, respectively). It seems that the geographical origin of H. officinalis L. greatly influences the oil quality. The essential oil of H. officinalis L. plant has been widely studied in Iran and other countries but the chemical composition of the essential oil of H. officinalis grown in Esfahan province is yet to be determined.

 The results of present study showed that the major oil constituents of the flowers and leaves of H. officinalis L. from Esfahan province, Iran were beta-pinene, trans-Pinocamphone and Cis-pinocamphone. In 1997, Veres et al. found that the oils from nine collections of H. officinalis grown from seed of different sources could be categorized depending upon their percentage composition of beta-pinene, limonene, pinocamphone and isopinocamphone. The oils were rich in isopinocamphone (5-50%), pinocamhone (3-50%) or contained beta-pinene and limonene (1-60%) as major the components. According to the ISO 9847/1991 standard, commercial oil should contain 40-67.5 % monoterpene ketones and 13.5-23.0 % β-pinene (Mazzanti et al. 1998). In this study, the values obtained for flowers were 46.83 % and 23.38 % for monoterpene ketones β-pinene in flowers and 43.6 % and 31.2 % β-pinene in leaves respectively. Figueredo et al. (2012) revealed that the major constituents of Hyssopus officinalis grown in Turkey were pinocarvone (29.2 %), trans-Pinocamphone (27.2 %), β-pinene (17.6 %), cis-pinocamphone (4.7 %) and myrcene (2.92 %). The literature data show that cispinocamphone compound can be predominate in hyssop oil, simultaneously with a low content of trans-pinocamphone (Mazzanti et al. 1998, Baj et al. 2010, Wesołowska et al. 2010). According to this, the examined sample belong to oils rich in β-pinene, cis-pinocamphone and transpinocamphone. Our results were in accordance with the most of the previously published. Cis-pinocamphone and transpinocamphone were the dominant constituents in hyssop oil in the studies of Mitić and Dordević (2000), Fraternale et al. (2004), Rosłon et al. (2002), Rey et al. (2004), and Zheljazkov et al. (2012). Mitić and Dordević (2000) showed that the content of cispinocamphone was at a level of 44.7%, whereas the content of trans–pinocamphone were lower (14.1%) than the present study. In this study, a higher amounts of β-pinene was found in flowers and leaves (23.38% and 31.2%, respectively). The oils from hyssop plants grown in Italy (Mazzanti et al. 1998, Fraternale et al., 2004), India (Garg et al., 1999), Egypt (Salma., 2002) and Hungary (Rey et al., 2004) were rich in β-pinene.

The other ther important compounds which were indentied in our study were β-ocimene, limonene, α-phellandrene, gamma-terpinene. The components with small amount were β-gurjunene, myrtenol, α-terpinolene, β-caryophyllene, linalool and camphene. Furthermore, 0.33% and 0.2% of in the were determined oleic acid flowers and leaves, respectively.

 

Conclusion

 

Our results demonstrated that in the flowering stage of officinalis L. of the major constituents grown in Isafahan essential oil from the flowers and leaves, respectively were β-pinene (23.38% and, 31.2%), trans-Pinocamphone (19.34%, 15.5%), cis-pinocamphone (27.49%, 28.1%), myrcene (6.01%, 6.0 %) and Sabinene (5.89%, 6.4%).

 

References

Benedec D, Oniga I, Tipercius B, Popescu H. 2002. Preliminary research of some polyphenolic compounds from Hyssopus officinalis L. (Lamiaceae). Farmacia, 50: 54-57.

Baj T, Kowalski R, Świątek Ł, Modzelewska M, Wolski T.  2010. Chemical composition and antioxidant activity of the essentials oil of hyssop (Hyssopus officinalis L.). Annales UMCS Sec. DDD, Pharmacia 3, 7: 55–62.

Benhammou N, Bekkara F A, Panovska TK. 2008. Antioxidant and antimicrobial activities of the Pistacia lentiscus and Pistacia atlantica extracts. African Journal of Pharmacy and Pharmacology, 2: 022-028.

Dragland S, Senoo H, Wake K, Holte K, Blomhoff  R .2003. Several culinary and medicinal herbs are important sources of dietary antioxidants. The Journal of Nutrition, 133: 1286-1290.

De Martino L, De Feo V, Nazzaro F.  2009. Chemical composition and in vitro antimicrobial and mutagenic activities of seven Lamiaceae essential oils. Molecules, 14: 4213-4230.

Fraternale, D, Ricci, D, Epifano  F, Curini, M. 2004. Composition and antifungal activity of two essential oils of hyssop (Hyssopus officinalis L.). Journal of Essential Oil Research, 16(6): 617-622.

Figueredo G, Özcan MM, Chalchat JC, Bagci Y, Chalard P.  2012.Chemical composition of essential oil of  Hyssopus officinalis L. and Origanum acutidens. Journal of Essential Oil Bearing Plants, 15 (2): 300 – 306.

Gollapudi S, Sharma HA, Aggarwal S, Byers LD, Ensley HE, Gupta S. 1995. Isolation of a previously unidentified polysaccharide (MAR-10) from Hyssopus officinalis that exhibits strong activity against human immunodeficiency virus type 1. Biochemical and Biophysical Research Communications, 210: 145-151.

Grzeszczuk  M, Jadczak  D.  2009. The estimation of biological value of some species of spice herbs. Acta Horticulturae, 830: 681-6.

Garg SN, Naqvi AA, Singh A, Ram G, Kumar S. 1999. Composition of essential oil from an annual crop of Hyssopus officinalis grown in Indian plains. Flavour and Fragrance Journal, 14: 170-172.

Ghalem BR, Mohamed B. 2009. Antimicrobial activity evaluation of the oleoresin oil of Pistacia vera L. African Journal of Pharmacy and Pharmacology, 3: 092-096.

Hornok, L. 1992. Cultivation and Processing of Medicinal Plants, John Wiley and Sons, Chichester, UK.

Hold KM, Sirisoma NS. 2002. Metabolism and mode of action of cis and trans-3-pinanones (the active ingredients of hyssop oil). Xenobiotica, 32: 251-265.

Jung EJ, Hyeok KS, Dong LG, Sang LY .2004. Production method of Hyssopus officinalis L. beverage. R. K. K. T. Kongbo. Korea. KR 2004013528.

Kizil S, Toncer O, Ipek A, Arslan N, Saglam S, Khawar KM. 2008. Blooming stages of Turkish hyssop (Hyssopus officinalis L.) affect essential oil composition. Acta  Agriculturae Scandinavica, Section B-Soil and Plant Science, 58(3): 273-279.

Kazazi H, Rezaei K, Ghotbsharif S, Emamdjomeh Z, Yamini Y. 2007.  Supercriticial fluid extraction of flavors and fragrances from Hyssopus officinalis L. cultivated in Iran. Food Chemistry, 105: 805-811.

Lugasi A, Hovari J, Hagymasi K, Jakoczi I, Blazovics A. 2006. Antioxidant properties of a mixture of Lamiaceae plants intended to use as a food additive. Acta Alimentaria, 35: 85-97.

Lawless J .2002. The Encyclopedia of Essential Oils. Thorsons, 110-11.

Mitić V. Đorđević S. 2000. Essential oil composition of Hyssopus officinalis L. cultivated in Serbia. Fact Universities. Series: Physics, Chemistry and Technology Vol. 2, No 2, pp. 105 – 108.

Michalczyk, M, Macura, R, Tesarowicz, I, Banaś, J. 2012. Effect of adding essential oils of coriander (Coriandrum sativum L) and hyssop (Hyssopus officinalis L.) on the shelf life of ground beef. Meat Science, 90(3): 842-850.

Mazzanti G, Battinelli L, Salvatore G. 1998. Antimicrobial properties of the linalool-rich essential oil of Hyssopus officinalis L var. decumbens (Lamiaceae). Flavour and Fragrance Journal, 13: 289–294.

Ozer H, Sahin F, Kilic H, Gulluce M. 2005. Essential oil composition of Hyssopus officinalis L. subsp. Angustifolius (Bieb.) Arcangeli from Turkey. Flavour and Fragrance Journal, 20:42 44.

Ozer H, Fikrettin S, Kilic H, Gulluce M. 2005. Essential oil composition of Hyssopus offcinalis L. subsp. angustifolius (Bieb.) Arcangeli from Turkey. Flavour and Fragrance Journal, 20: 42-44.

Rey C, Carron CA, Cottagnoud A, Bruttin B, Carlen C. 2004. The hyssop (Hyssopus officinalis) cultivar ‘Perlay’. Revue suisse de viticulture arboriculture horticulture, 36(6):337–341.

Rosłon W, Osińska E, Węglarz Z. 2002. Evaluation of three species of Hyssopus genus with respect to their development as well as essentials oil content and its composition. Folia Horticulturae, 14(2): 145–151.

Shah NC, Kahol AP, Sen T, Uniyal GC. 1986. Gas chromatographic examination of oil of Hyssopus officinalis. Parfuemerie und Kosmetik, 67: 116, 118.

Salma AS. 2002. Chemical and physiological studies on anise hyssop (Agastache foeniculum Pursh) and hyssop (Hyssopus officinalis L) plants grown in Egypt as new spices. Bulletin of the National Research Centre, 27: 25-35.

Veres  K, Varga  E, Dobos A, Hajdn  Z, Mathe  I, Pluhar  Z, Nemeth  E, Bernath J. 1997. Investigation of thecomposition of essential oils of Hyssopus officinalis L. populations, 217-220 pp. In: Ch. Franz, A. Mathe, G. Buchbaner (Eds), Essential Oils: Basic and Applied Research, AlluredPublishing Corporation, Carol Stream, IL, pp. 217220.

Varga E, Hajdu Z, Veres K, Mathe I, Nemeth E, Pluhar Z, Bernath J. 1998a. Investigation of variation of the production of biological and chemical compounds of Hyssopus officinalis L. Acta Pharmaceutica Hungarica, 68: 183-188.

Wesołowska A, Jadczak D, Grzeszczuk M. 2010. Essential oil composition of hyssop (Hyssopus officinalis L.)

Zheljazkov VD, Astetkie T, Hristov AN. 2012. Lavender and hyssop productivity, oil content, and bioactivity as a function of harvest time and drying. Industrial Crops and Products, 36: 222–228.

Zhao, J. Davis, LT, Verpoort, R. 2005. Elicitor signal transduction leading to production of plant secondary metabolites. Biotechnology Advances. 23: 283–333.

Zargari A. 1992. Medicinal plants. Tehran University Press, Tehran, Iran.