Lipid Fatty Acids and Essential Oil from Ferula prangifolia Endemic in Uzbekistan

Lipids and essential oils from Ferula prangifolia Korovin (Apiaceae) endemic in Uzbekistan were studied for the first time. The compositions of lipid fatty acids and essential oils of fruit and leaves were established.

Keywords: Ferula prangifolia Korovin; lipids; petroselic acid; hexadecatrienoic acid; essential oil; GC-MS; GC-FID

Translated from Khimiya Prirodnykh Soedinenii, No. 2, March–April, 2022, pp. 190–194.

The genus Ferula L. comprises many (22%) of the greater than 200 species of plants in the family Apiaceae (order Apiales) in the flora of Uzbekistan. Ferula species, like most medicinal plants in this family, contain coumarins, flavonoids, resins, essential oils, and other biologically active compounds. These plants bolster the number of traditional medicinal agents of Eastern medicine because of their variety of chemical compositions and biological activities [[1]–[3]]. Essential oils of several Ferula species growing in Kazakhstan have been studied. Changes in their contents and chemical compositions as a function of plant organ and isolation method were reported [[4]–[6]].

Ferula prangifolia Korovin is one of eight Ferula species endemic in the flora of Uzbekistan. Information on lipids and volatile compounds of this plant has not appeared in the scientific literature except for our short publication on the fruit [[7]]. Previously, we reported the compositions of essential oil and lipids from leaves of another endemic species, F. kuhistanica Korovin [[8]].

The goal of the present research was to compare fatty acids of lipids and essential oils from fruit and leaves of F. prangifolia. Lipids, fatty acids, and essential oil (EO) were analyzed by traditional methods under conditions analogous to those described before [[8]]. Table 1 lists the contents of EO, lipid groups, lipophilic substances (LS), and total lipids (TL) in fruit and leaves of the plant.

Table 1 Contents of Essential Oils and Lipids in Fruit and Leaves of F. prangifolia, mass% of Dry Substance

Plant organ	EO	NL	PL	TL	LS*
Fruit	0.7	11.8 (91.5)*	1.1 (8.5)	12.9	1.5
Leaves	0.5	3.3 (51.6)	3.1 (48.4)	6.4	14.8

*Fraction of lipophilic substances, NL, and PL in TL, %

Table 1 shows that the yield of EO from fruit of F. prangifolia was 1.4 times greater than from leaves. Fruit contained a moderate amount of oil (12.9%) with the neutral lipid (NL) fraction making up 91.5% of it. Leaves held a sightly greater level of NL (51.6%) than polar lipids (PL, 48.4%). Lipids isolated from leaves were more enriched in LS than those from fruit.

TLC analysis of lipids in extracts of fruit and leaves found that they consisted of acyl-containing lipids (triacylglycerins, fatty alcohol and sterol esters, glyco- and phospholipids), free fatty acids, and lipophilic compounds. The major constituents of fruit TL were triacylglycerins (94.7%).

Leaf lipids also contained chlorophyll pigments although galactolipids and free fatty acids dominated the separate constituents. Leaf galactolipids were represented by mono- and digalactosyldiacylglycerins. Carotenoids, hydrocarbons, fatty alcohols, triterpenols, sterols, and essential oil components were detected in LS from fruit and leaves.

Table 2 indicates that unsaturated fatty acids were the predominant acyl fragments in TL constituents from fruit and leaves (87.6 and 89.0%, respectively). The main fraction in fruit consisted of linoleic (18:2ω6, 47.1%) and isomeric octadecenoic (39.3%) oleic (18:1ω9) and petroselic acids (18:1ω12).

Table 2 Composition of Lipid Fatty Acids of Fruit and Leaves of F. prangifolia, % by GC

Saturated fatty acid	Fruit	Leaves	Unsaturated fatty acid	Fruit	Leaves
12:0	0.1	0.4	16:1ω7	0.7	0.3
14:0	0.6	0.8	16:3ω3	–	0.4
15:0	0.2	0.1	18:1ω9	39.3	19.7
16:0	7.9	7.1	18:1ω12		–
17:0	Tr.	0.1	18:2ω6	47.1	19.3
18:0	2.9	1.9	18:3ω3	Tr.	49.0
20:0	0.7	0.4	20:1ω11	0.5	0.3
22:0	Tr.	0.2	Total	87.6	89.0
Total	12.4	11.0

Leaf lipids were dominated by linolenic acid (18:3ω3). Also, small amounts of (Z,Z,Z)-hexadeca-7,10,13-trienoic acid (16:3ω3, 0.4%) were detected in them. The content of this specific acid was significantly greater (9.4%) in leaves of the previously studied species F. kuhistanica. Therefore, F. prangifolia belonged to the 18:3-plant group, in contrast to the related endemic taxon F. kuhistanica (16:3 plant) [[8]].

EOs isolated from fruit and leaves of F. prangifolia were analyzed by GC over nonpolar and polar columns using MS and FID detectors. The results indicated that the main constituents of the volatile compounds from fruit were the sesquiterpenes δ-cadinene, bicyclogermacrene, α-muurolene, γ-cadinene, β-elemene, and their oxygenated derivatives such as α-cadinol, T-muurolol, and T-cadinol (41.6 and 47.1%, respectively) (Table 3). The sesquiterpenes were dominated by α-cadinol. Oxygenated sesquiterpenes with a high content (18.3%) of spathulenol made up almost one third of EO from leaves. Next in content were constituents such as thymol, carvacrol, trans-α-bergamotol, T-cadinol, caryophyllene oxide, and pinenes. The composition of volatile compounds from leaves featured a high level of fatty acids 16:0 and 14:0 (total 31%).

Table 3 Composition of Essential Oils from Fruit and Leaves of F. prangifolia, GC-MS

Compound	RRIa	RRIb	Fruit		Leaves
			HP-5, % by MS	HP-INNOWax, % by FID
Hexanal	803	1093			0.8
Nonane	900	900			Tr.
α-Thujene	928	1035	0.4	Tr.	Tr.
α-Pinene	934	1032	1.4	0.5	1.5
Camphene	950	1076	0.1		Tr.
2-Methylnonane	961	961	0.1		5.8
Sabinene	973	1132	0.1		Tr.
β-Pinene	976	1118	0.1		1.5
Mycene	990	1174	0.3
α-Terpinene	1015	1188			Tr.
Decane	1000	1000	0.2
δ-3-Carene	1009	1159	Tr.
para-Cymene	1023	1280	0.6	0.3	Tr.
Limonene	1027	1203	1.0	0.5	0.9
(E)-β-Ocimene	1036	1246	0.1
γ-Terpinene	1058	1255			Tr.
α-Methyldecane	1061	1065	0.9	0.7
para-Cymenene	1090		0.2
Linalool	1194	1553		Tr.
Undecane	1100	1100	1.3	1.0	0.9
Pinocarvone	1160	1586			Tr.
(E)-Pinocarveol	1138	1670	0.1
Terpinen-4-ol	1175	1611	0.3	Tr.	Tr.
Myrtenal	1196	1648			Tr.
(E)-Carveol	1218	1845	Tr.
Citronellol	1228	1770	0.4	Tr.
Thymol methyl ether	1232	1604	0.1	Tr.
Thymol	1293	2198	0.1		5.6
Carvacrol	1300	2239	0.1		3.9
Bicycloelemene	1334	1495	Tr.	Tr.
α-Cubebene	1347	1466	0.1	0.2
Ledene	1369	1708	Tr.	0.4
α-Copaene	1372	1497	0.7	0.5
2-epi-α-Funebrene	1382		0.4
β-Elemene	1391	1600	6.6	3.2	1.3
(Z)-Jasmone	1397	1969	0.1
β-Funebrene	1410	1594	Tr.	0.1
β-Caryophyllene	1416	1612	0.9	1.3	Tr.
β-Copaene	1426	1606	0.2
Calarene (= β-Gurjunene)	1430	1610	0.2
Aromadendrene	1435	1628	0.3	0.2
Pulegone		1662			Tr.
β-Barbatene	1438	1667	0.1
α-Humulene	1450	1687	0.4	0.6	Tr.
allo-Aromadendrene	1456	1661	0.2	Tr.
Verbenene		1725			Tr.
γ-Murrolene	1475	1704	1.2	1.3
Germacrene D	1479	1726	1.2	2.4
(E)-β-Ionone	1485	1958	1.0	0.2
Bicyclosesquiphellandrene	1489	1738	0.2	0.4
epi-Cubebol	1489	1900		0.2
β-Selinene	1490	1742			Tr.
α-Selinene	1498	1744	3.2	0.4	Tr.
Bicyclogermacrene	1498	1753		7.6	Tr.
α-Muurolene	1499	1740		4.8	Tr.
epi-Zonarene	1499	1677	3.0	Tr.	Tr.
Cuparene	1499	1849		Tr.	Tr.
(E)-Verbenol		1683			Tr.
β-Bisabolene	1506	1741	0.8		Tr.
γ-Cadinene	1513	1776	2.9	3.7	0.4
Cubebol	1513	1957		0.2
(E)-Calamenene	1523	1853	5.9	0.8
δ-Cadinene	1523	1773		13.9	0.9
Cubenene (=Cadina-1,4-diene)	1535	1799		0.8
Citronellyl butyrate	1535	1786	0.5	0.4
α-Cadinene	1535	1743		0.8
Diisobutyl maleate*		1909			1.5
α-Calacorene II	1540	1942	0.2	0.4
Selina-4,11-diene	1549	1688	0.3	Tr.
β-Calacorene	1560	1918	0.1	0.2
Germacrene D-4-ol	1571	2069	1.3	0.7
Spathulenol	1579	2144	7.7	4.1	18.3
Caryophyllene oxide	1580	2008		0.2	1.6
Globulol	1582	2098	2.4	1.0
Viridiflorol	1590	2104	1.1	Tr.
Cubeban-11-ol	1599	2080	0.2	Tr.
Rosifoliol	1599	2143		Tr.
β-Oplopenone	1604	2092	0.3		0.7
Humulene epoxide II		2010			0.4
1,10-Diepicubenol	1610	2081	0.5
α-Corocalene	1618	–	0.1
1-Epicubenol	1624	2088	0.5	1.0
Linalool ether*		2161	–	4.0
T-Muurolol	1643	2209		7.5
Torreol (=α-Muurolol, δ-Cadinol)	1643	2219		2.6
T-Cadinol	1643	2187	10.6	4.6	1.8
Cubenol	1643	2080		1.1
β-Eudesmol	1658	2257		0.4	1.6
α-Eudesmol	1658	2250			0.4
α-Cadinol	1658	2255	22.4	20.8	1.3
Selin-11-en-4-α-ol	1658	2273			0.4
10-Hydroxycalamenene	1658	2290		0.4
(E)-α-Bergamotol	1684	2247	0.6	1.4	3.1
(Z,E)-α-Bergamotol	1720	2328	0.3	0.3
(Z)-Lanceol	1759	2342	2.4	0.4
Tetradecanoic acid		2670			2.0
Hexahydrofarnesyl acetone	1839	2131	0.2	0.1	1.3
Farnesyl acetone		2248			Tr.
Hexadecanoic acid		2931		Tr.	29.0
Phytol	2110	2622		1.0
Nonacosane	2900	2900			2.6
Sum of identified compounds, of them:			91.1	99.2	89.5
monoterpene hydrocarbons			4.2	1.3	3.9
oxygenated monoterpenes			3.0	0.6	9.5
sesquiterpene hydrocarbons			29.7	41.6	2.6
oxygenated sesquiterpenes			50.4	47.1	30.9
fatty acids			–	Tr.	31.0
others			3.7	8.6	46.9

aRRI, Relative retention index of constituent on HP-5 column; bRRI, relative retention index of constituent on HP-INNOWax column; Tr, trace > 0.1%; *preliminary identification based on comparison of mass spectra.

Thus, comparisons of fatty acids of lipids and EOs from F. prangifolia using chromatographic columns of different polarities found that the main constituents of fatty oil from fruit were triacylglycerins and esterified linoleic and isomeric octadecenoic acids (6Z and 9Z); from leaves, (Z,Z,Z)-hexadeca-7,10,13-trienoic acid. EO constituents of fruit and leaves were dominated by α-cadinol and spathulenol, respectively.

Experimental

GC-MS analysis of EO used an Agilent 5975 GC-MSD chromatograph (Agilent, USA; SEM Ltd., Istanbul, Turkey) equipped with a polar HP-INNOWax FSC column (60 m × 0.25 mm × 0.25 μm; Agilent, USA). The chromatography temperature (60°C) was held for the first 10 min then raised to 220°C at 4°C/min, holding at that temperature for 10 min, after which it was raised to 240°C at 1°C/min and held there for 35 min. The total analysis time was 115 min. Mass spectra were scanned at ionizing-electron energy 70 eV. The mass range covered m/z from 35 to 450 amu. GC-FID analysis used an Agilent 6890N GC chromatograph (SEM Ltd., Istanbul, Turkey) and identical conditions using an analogous column and a flame-ionization detector (FID). The other chromatography conditions were published before [[8]].

In parallel, EO were chromatographed on an Agilent 7890A GC/5975C Inert MSD instrument (Agilent Technologies, USA) over an HP-5ms column (30.0 m × 250.0 μm × 0.25 μm). The temperature was held at 70°C for 4 min, increased to 250°C at 4°C/min, held there for 6 min, and then raised to 270°C at 10°C/min. The injector temperature was 280°C. The mass spectral conditions were ionization energy 70 eV, source temperature 180°C, quadrupole 150°C, mass range 10–500 amu. Constituents were identified by comparing mass spectra to data in electronic libraries (Wiley Registry of Mass Spectral Data, 9th Ed.; NIST Mass Spectral Library, 2011, W9N11.L; Baser Library of Essential Oil Constituents; and Adam′s Library) and by the relative retention index (RRI) of compounds that was determined relative to retention times of a mixture of C9–C24n-alkanes. The MassFinder 3.0, Wiley GC-MS Library was used.

Fatty acid methyl esters were chromatographed on an Agilent 6890N instrument equipped with an FID. An HP-5 capillary column (30 m × 0.32 mm) was used. The column temperature was held at 70°C for 4 min, raised to 270°C at 10°C/min, and held there for 10 min. The carrier gas was He for all chromatographs. The models for GC-MS and GC-FID were fatty acid methyl esters prepared earlier from lipids of F. kuhistanica leaves and Nigella sativa seeds [[8]]. Constituents were identified based on RRI values [[9]] and TLC on silica gel with added AgNO3 (20%) in C6H6 [[10]]. Oleic and linolenic acids were quantitatively analyzed using GC-MS data [on an Agilent 7890A GC/5975C Inert MSD instrument with an HP-5ms column (30.0 m × 250.0 μm × 0.25 μm)] from the intensities of the molecular masses of their methyl esters (m/z 296 and 292, respectively).

The plant was collected in 2013 in Chakak District, Tashkent Region (Uzbekistan) during flowering; fruit, during fruiting. An herbarium specimen of the plant is preserved in the collection of the Laboratory of Medicinal and Technical Plants, AS, RUz, under number 1364.

EOs from F. prangifolia fruit and leaves were isolated by steam distillation in a Clevenger apparatus according to the pharmacopoeial method [[11]].

Fatty acids were isolated and analyzed by first obtaining TL via extraction of ground plant organs by the Folch method using a mixture of CHCl3 and MeOH (2:1). The purification and fractionation conditions of the TL by column chromatography, analysis of NL and PL constituents using TLC, isolation of LS and fatty acids from TL by alkaline hydrolysis, and methylation of acids by diazomethane have been described in detail [[8], [12]].

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By D. T. Asilbekova; G. Ozek; T. Ozek; Kh. M. Bobakulov; A. M. Nigmatullaev and Sh. Sh. Sagdullaev

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