Sulphur-containing compounds in the essential oil of Ferula alliacea roots and their mass spectral fragmentation patterns

Abstract

Context GC-MS analysis is the best way to characterize volatile sulphur-containing compounds. Ferula (Apiaceae) is a genus of perennial herbs. Due to the occurrence of essential oils or oleoresins in the Ferula species, these plants usually possess strong aromatic scent. Terpenoid compounds were the most abundant constituents of Ferula oils, however, in some of Ferula species, the essential oils were dominated by volatile sulphur-containing compounds.

Objectives Ferula alliacea Boiss. is considered one of the sources of the oleo-gum-resin asafoetida. In this study, we analyzed the hydrodistilled essential oil from its dried roots and provide new data about retention indices and mass fragmentation patterns of some volatile sulphur-containing compounds that are useful for future studies on this class of compounds.

Materials and methods The roots of F. alliacea were collected during the flowering stage of plant, from Bezgh, Kashmar to Neishabour road, Khorasan-Razavi province, Iran, in June 2012. The oil was obtained by hydrodistillation using a Clevenger apparatus and analyzed by GC-MS.

Results This is the first report on phytochemical analysis of F. alliacea roots. Seventy-six components, representing 99.5% of the oil, were characterized. The major components were 10-epi-γ-eudesmol (22.3%), valerianol (12.5%), hinesol (8.3%), guaiol (7.3%) and Z-propenyl-sec-butyl trisulphide (6.5%). Predominant mass fragment ions of the identified sulphur-containing compounds are explained in this paper.

Conclusion The volatile oil of F. alliacea mostly contains oxygenated sesquiterpenes, however, its odour was dominated by sulphur-containing compounds. The most abundant sulphur-containing compound includes Z-propenyl-sec-butyl trisulphide (6.5%).

Keywords:

• Apiaceae
• mintsulphide
• oleo-gum-resin asafoetida
• oxygenated sesquiterpenes

Introduction

Ferula (Apiaceae), a genus of perennial herbs, comprises about 170 species distributed from central Asia (particularly Iran and Afghanistan) westward throughout the Mediterranean region to northern Africa (Pimenov & Leonov 1993). Thirty species of Ferula have been represented in Iranian flora, of which 15 species are endemic to Iran (Mozaffarian 1983, 1996). To date, more than 70 species of this genus have been chemically investigated (Iranshahi et al. 2008c, 2010a,b; Sahebkar & Iranshahi 2011; Kasaian et al. 2014). Based on the findings, Ferula spp. have been turned out to be a source of bioactive phytochemicals such as sesquiterpene/sesquiterpene coumarin derivatives (Iranshahi et al. 2004, 2007, 2008d), and sulphur-containing compounds (Iranshahi et al. 2003, 2006, 2008b; Sahebkar et al. 2010; Iranshahi 2012).

The most important medicinal Ferula species include F. assa-foetida Boiss. & Buhse, F. gummosa Boiss. F. foetida Regel, F. latisecta Rech. f. & Aellen and F. persica Willd. Previous research also unveiled a number of medicinal properties form Ferula spp. including antibacterial, antifungal, antioxidant, anti-inflammatory, antinociceptive, anticonvulsant, antispasmodic and hypotensive activities (Bakkali et al. 2008; Asili et al. 2009; Maggi et al. 2009; Bagheri et al. 2010; Franz 2010; Chibani et al. 2012; Labed-Zouad et al. 2015).

Volatile sulphur compounds have been reported from different terrestrial plants (Iranshahi, 2012). One of the most well-known volatile sulphur compounds is allicin from Allium species. A large number of biological activities have been attributed to allicin and its analogues including antibacterial, antifungal, antiviral and antiprotozoal properties (Borlinghaus et al. 2014). However, the scaffold of volatile sulphur-containing compounds of Ferula species is unique and has only been reported from the genus Ferula (Iranshahi 2012). There are a few reports on the biological activities of volatile sulphur compounds from Ferula species such as their antifungal property against human dermatophytes (Iranshahi et al. 2008a).

Ferula alliacea Bioss. is used in Iranian folklore as one of asafoetida sources with similar medicinal applications to those of F. assa-foetida. This plant possessed a strong sulphurous odour due to the presence of volatile sulphur-containing compounds. No phytochemical investigation has been conducted on the volatile sulphur compounds of this species. In this study, we aimed to identify chemical composition of the essential oil of F. alliacea roots for the first time. The spectral mass fragmentation patterns of sulphur-containing compounds are also discussed in this paper. These mass fragmentation patterns can provide useful information for researchers who are interested in identifying the volatile sulphur-containing compounds of this genus.

Materials and methods

Plant material

The roots of F. alliacea were collected during the flowering stage of plant, from Bezgh, Kashmar to Neishabour road, Khorasan-Razavi province, Iran, in June 2012. The plant was identified by Mr. Joharchi and a voucher specimen (No. 38927) was deposited at the Herbarium of the Department of Pharmacognosy, School of Pharmacy, Mashhad University of Medical Sciences, Mashhad, Iran.

Essential oil preparation

Essential oil (EO) was isolated using hydro-distillation of air-dried material for 3 h, with a Clevenger-type apparatus. The oil was obtained as a greenish yellow liquid with a yield of 0.13% (v/w). The oil was dried over anhydrous sodium sulphate and stored at 4 °C in dark until use. The EO was dissolved in n-hexane for gas chromatography and mass spectrometry analyses.

GC-MS analysis

The GC-MS analyses were performed using an Agilent 5975 apparatus (Agilent Technologies, Santa Clara, CA) with a HP-5 MS column (30 m × 0.25 mm i.d., 0.25 m film thicknesses) interfaced with a quadruple mass detector and a computer equipped with Wiley 7n. Library. Other analytical settings were: oven temperature: 50 °C (5 min), 50–250 °C (3 °C/min), 250 °C (10 min); injector temperature 250 °C; injection volume: 0.1 L; split ratio: 1:50; carrier gas: helium at 1.1 mL/min; ionization potential: 70 eV; ionization current: 150 A; and mass range: 35–465 mui. Identification of individual compounds was made by comparison of their mass spectra and retention indices (RI) with those of authentic samples and those given in the literature (Khajeh et al. 2005; Adams 2007). Quantification of the relative amount of each individual component was performed according to the area percentage method without consideration of calibration factor.

Results and discussion

The oil of the roots of Ferula alliacea was greenish yellow, in a total yield of 0.13% v/w. Seventy-six components were identified from the oil of this species (Table 1), representing 99.5% of the total oil. GC-MS analysis of the essential oil of F. alliacea roots led to the identification of mono- and sesquiterpenes as well as volatile sulphur-containing compounds. Although oxygenated sesquiterpenes were determined to be the most abundant constituents in the essential oil, the odour of the oil was dominated by volatile sulphur-containing compounds. The essential oil contained oxygenated sesquiterpenes (74.7%), sulphur-containing compounds (16.6%), sesquiterpene hydrocarbons (3.7%) and oxygenated monoterpenes (2.6%). The major components of the oil were determined to be 10-epi-γ-eudesmol (22.3%), valerianol (12.5%), hinesol (8.3%), guaiol (7.3%) and Z-propenyl-sec-butyl trisulphide (6.5%). The sulphur-containing compounds (16.6% of the oil, Table 2) were identified to be 3,4-dimethyl thiophene (trace), 2,3,4-trimethyl thiophene (trace), propyl sec-butyl disulphide (0.1%), Z-1-propenyl sec-butyl disulphide (2.2%), E-1-propenyl sec-butyl disulphide (1.9%), di-sec-butyl disulphide (0.1%), methyl 1-(methylthio) propyl disulphide (0.1%), Z-1-propenyl propyl trisulphide (0.3%), E-1-propenyl propyl trisulphide (0.4%), Z-1-propenyl sec-butyl trisulphide (6.5%), E-1-propenyl sec-butyl trisulphide (2.7%), sec-butenyl sec-butyl trisulphide (0.1%) and mintsulphide (0.3%). The volatile sulphur-containing compounds were characterized on the basis of their retention indices (Iranshahi 2012) and mass spectral fragmentation patterns (Figure 1 and Table 2).

Figure 1. Major mass fragmentation ions of the volatile sulphur-containing compounds from Ferula alliacea.

Table 1. Chemical composition of the essential oil from the roots of Ferula alliacea Boiss.

NO	Compound	RI^a	Percentage
1	3,4-Dimethyl thiophene	905	t^b
2	Methyl sec-butyl disulphide	994^c	t
3	2,3,4-Trimethyl thiophene	1015	0.1
4	trans-Verbenol	1149	t
5	n-Propyl-sec-butyl disulphide	1167	0.1
6	Borneol	1169	0.1
7	Z-1-Propenyl-sec-butyl disulphide	1171	2.2
8	Lavandulol	1174	0.1
9	E-1-Propenyl-sec-butyl disulphide	1176	1.9
10	iso-Dihydrocarveol	1203	0.5
11	di-sec-Butyl disulphide	1214	0.1
12	endo-Fenchyl acetate	1224	0.1
13	Nerol	1234	0.1
14	Carvone	1248	t
15	Methyl-1-(methylthio) propyl disulphide	1259	0.1
16	Bornyl acetate	1290	0.1
17	Lavandulyl acetate	1297	0.3
18	trans-Pinocarvyl acetate	1303	t
19	Terpinen-4-ol acetate	1306	0.3
20	δ-Terpenyl acetate	1328	0.8
21	Z-Propenyl-propyl trisulphide	1349	0.3
22	E-Propenyl-propyl trisulphide	1353	0.3
23	Neryl acetate	1373	0.1
24	α-Copaene	1379	0.1
25	n-Propyl sec-butenyl trisulphide	1393	0.4
26	β-Elemene	1395	0.3
27	β-Funebrene	1415	0.1
28	trans-Caryophyllene	1422	0.2
29	Z-Propenyl-sec-butyl trisulphide	1433	6.5
30	E-Propenyl-sec-butyl trisulphide	1435	2.7
31	β-Barbatene	1445	0.2
32	α-Humulene	1457	0.2
33	allo-Aromadendrene	1461	0.1
34	β-Farnesene	1465	0.1
35	Inseparable mixture of E and Z-propenyl-1-(methylthio) propyl disulphide	1473	1.0
36	γ-Murrolene	1481	0.2
37	α-Amorphene	1484	t
38	β-Chamigrene	1487	0.1
39	β-Selinene	1489	0.3
40	δ-Selinene	1496	0.2
41	Bicyclogermacrene	1499	0.5
42	β-Himachalene	1502	0.8
43	β-Dihydroagarofurane	1506	0.3
44	Cuparene	1509	0.3
45	sec-Butenyl-sec-butyl trisulphide	1511	0.1
46	(Z)-γ-Bisabolene	1515	0.2
47	Butylated hydroxy toluene	1519	0.6
48	Myristicin	1528	0.7
49	α-Agarofuran	1549	1.2
50	Elemol	1555	0.8
51	Elemicin	1565	0.4
52	E-Nerolidol	1572	0.2
53	Spathulenol	1582	0.3
54	Viridiflorol	1595	0.2
55	Guaiol	1606	7.3
56	5-epi-7-epi-α-Eudesmol	1615	4.7
57	10-epi-γ-Eudesmol	1630	22.3
58	γ-Eudesmol	1632	0.2
59	Eremoligenol	1638	2.0
60	Hinesol	1645	8.3
61	Agarospirol	1647	0.9
62	α-Eudesmol	1658	3.6
63	Valerianol	1663	12.5
64	7-epi-α-Eudesmol	1666	2.4
65	(2Z-6Z)-Farnesal	1680	2.3
66	α-epi-Bisabolol	1691	0.1
67	α-Bisabolol	1693	0.4
68	Inseparable mixture of E and Z-propenyl 1-(methylthio) propyl trisulphide	1699	0.4
69	(2Z-6Z)-Farnesol	1707	0.2
70	Mintsulphide	1741	0.3
71	Unknown	1749	0.2
72	Cedryl acetate	1768	0.4
73	Hinesol acetate	1784	1.4
74	α-Eudesmol acetate	1793	0.9
75	(2Z–6Z)-Farnesyl acetate	1800	1.8
76	Unknown	1822	0.3
	Major grouped compounds
	Monoterpene hydrocarbons		0
	Oxygenated monoterpenes		2.6
	Sesquiterpene hydrocarbons		3.9
	Oxygenated sesquiterpenes		74.7
	Sulphur-containing compounds		16.6
	Miscellaneous compounds		1.7
	Total identified		99.5

^aRI: The Kovats retention indices relative to C8–C20 n-alkanes were determined on CP-Sil 8CB capillary column.

^bt: trace > 0.05%.

^cFor the RIs of sulphur-containing compounds, see Iranshahy and Iranshahi (2011), Sahebkar et al. (2010), and Sahebkar and Iranshahi (2010).

Table 2. Mass fragmentation ions observed for volatile sulphur-containing compounds from the essential oil of Ferula alliacea roots. Predominant ions have shown in bold.

No.	Compound name	Mass fragmentation ions (percentage)
1	3,4-Dimethyl thiophene	112 (71), 111 (100), 97 (46), 77 (10), 69 (7), 45 (12)
2	Methyl sec-butyl disulphide	136 (63), 80 (100), 64 (9), 57 (32), 41 (43)
3	2,3,4-Trimethyl thiophene	126 (73), 125 (63), 111 (100), 97 (6), 91 (10), 77 (10), 59 (9), 45 (10)
5	n-Propyl sec-butyl disulphide	164 (57), 108 (100), 57 (50), 43 (78), 41 (64)
7	Z-1-Propenyl sec-butyl disulphide	162 (53), 106 (100), 57 (23), 41 (38)
9	E-1-Propenyl sec-butyl disulphide	162 (50), 106 (100), 57 (20), 41 (36)
11	di-sec-Butyl disulphide	178 (30), 122 (39), 57 (100), 41 (25)
15	Methyl 1-(methylthio) propyl disulphide	89 (100), 79 (13), 73 (13), 61 (21), 45 (24), 41 (40)
21	Z-Propenyl-propyl trisulphide	180 (3), 75 (100), 59 (10), 45 (10)
22	E-Propenyl-propyl trisulphide	180 (3), 75 (100), 59 (10), 45 (10)
25	n-Propyl sec-butenyl trisulphide	194 (3), 75 (100), 59 (10), 41 (11)
29	Z-Propenyl-sec-butyl trisulphide	194 (1), 89 (100), 73 (10), 61 (15), 45 (15), 41 (25)
30	E-Propenyl-sec-butyl trisulphide	194 (1), 89 (100), 73 (10), 61 (13), 45 (15), 41 (25)
35	Inseparable mixture of E and Z-propenyl 1-(methylthio) propyl disulphide	89 (100), 73 (7), 61 (10), 41 (21)
45	sec-Butenyl-sec-butyl trisulphide	208 (1), 89 (100), 73 (7), 61 (14), 41 (23)
68	Inseparable mixture of E and Z-propenyl 1-(methylthio) propyl trisulphide	89 (100), 73 (10), 61 (7), 57 (10), 41 (22)
70	Mintsulphide	236 (25), 159 (9), 123 (100), 112 (63), 91 (21), 79 (50), 67 (9), 55 (9), 41 (16).

In general, sec-butyl part of sulphur-containing compounds is considered as the chemotaxonomic marker of the family Apiaceae. sec-Butyl derivatives of sulphur-containing compounds have previously been reported from Ferula species including F. assa-foetida (Iranshahy & Iranshahi 2011), F. latisecta, (Iranshahi et al. 2008b; Sahebkar et al. 2010), F. persica (Iranshahi et al. 2003, 2006) and F. foetida (Chitsazian-Yazdi et al. 2015). As shown in Figure 1 and Table 2, the m/z = 89 ion peak is produced by the [sec-butyl-S] ion that could be formed by splitting the molecular ion at the SS or SSS bonds. This ion peak is usually observed as base peak. The m/z = 75 ion peak is also produced by the [propyl-S] ions and observed as a predominant peak in propyl-containing sulphur compounds. Considering thiophene derivatives, [M-H]⁺(111 and 125) and [M-CH₃]⁺(97 and 111) are predominant mass fragmentation ions in their mass spectra (Figure 1). Other common fragmentation ion peaks in the mass spectra of sulphur-containing compounds include 57, 41 and 43 attributable to [isobutyl], [allyl] and [propyl] ions, respectively.

In total, GC-MS analysis is the best approach to characterize sulphur-containing compounds. The characterization of these compounds is not commonly carried out using NMR techniques due but their small quantities and volatility. In this study, we tried to provide new information about the retention indices and mass fragmentation patterns of some volatile sulphur-containing compounds that are useful for future studies on this class of compounds.

The volatile oil of F. alliacea mostly contains oxygenated sesquiterpenes, however, its odour was dominated by sulphur-containing compounds. The most abundant sulphur-containing compound includes Z-propenyl-sec-butyl trisulphide (6.5%).

This plant is used by local people as an anthelminthic and antispasmodic herb for gastrointestinal disorders. There is a gap in our knowledge about the biological activities of sulphur components of F. alliacea and other Ferula species that needs further investigation; however, sulphur compounds may play a role, at least in part, in biological activities of this plant.

Funding information

This research was supported by Mashhad University of Medical Sciences (MUMS) Research Council.

Acknowledgements

The authors would like to thank Mr. H. Joharchi for his assistance in plant identification.

Disclosure statement

The authors report no declarations of interest.