Volatile Oil Composition of an Antiobesity Unani Formulation Safoof-E-Muhazzil

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Kamran Javed Naquvi et al. / Journal of Pharmacy Research 2012,5(1),12-15 Research Article Available online through ISSN: 0974-6943 http://jprsolutions.info Volatile oil composition of an antiobesity Unani formulation Safoof-e-Muhazzil

Kamran Javed Naquvi1, Shahid Husain Ansari1*, Mohammed Ali1, Abul Kalam Najmi2, Md Rafiul Haque1 1Department of Pharmacognosy and Phytochemistry, Faculty of Pharmacy, Jamia Hamdard, New Delhi-110062. India. 2Department of Pharmacology, Faculty of Pharmacy, Jamia Hamdard, New Delhi-110062. India. Received on:20-09-2011; Revised on: 15-10-2011; Accepted on:10-12-2011

ABSTRACT The hydro-distilled volatile oil of the Safoof-e-Muhazzil, an antiobesity Unani formulation, was analyzed using GC and GC-MS for the first time. Fifty eight compounds representing 99.35 % of the oil were characterized. Consisting of thirteen monoterpenes (11.62 %), forty four sesquiterpenes (85.05 %), two diterpene alcohols (0.56 %), trans-phytol and 13(16),14-labdiene-8-ol, four aliphatic hydrocarbons n-eicos-15-enol, 9,12-octadecadienoic acid methyl ester, n-heptacosane and methyl-6-hydroxyoctadecanoate and two aromatic components 2-benzilidine and amyrolin. Eudes-4(14),11-diene (28.61 %), viridiflorol laurate (16.40 %), bisabolene (9.73 %), globulol (9.13 %), thymol (6.14 %), t-cadinol (4.15 %), trans-cadine-1,4-diene (2.08 %), 2E, 6E-farnesol (1.41), L- limonene (1.39 %), -cadinene (1.37 %) and β-gurjunene (1.28) were the predominant compounds.

Keywords: Safoof-e-Muhazzil, volatile oil, GC-MS, sesquiterpenes, Unani formulation.

INTRODUCTION Sufoof-e-Mohazzil or Safoof Mohazzil, is one such polyherbal formulation ties. Lac is the only natural resin of animal origin, produced by a red coloured used by Unani physicians for treatment of obesity. The preparations mar- tiny Lac insect, Laccifer lacca (Kerr) found throughout India. Fourteen spe- keted by different manufacturers have minor differences in their composi- cies of insect Laccifer are reported in India on a number of plants both wild tion. The formulation given in National Formulary of Unani Medicine con- and cultivated in various regions. Host plants, viz. Scheichera oleosa (Lour.) tains Ajwain (Nankhwah-Trachyspermum ammi Linn., seeds) 10 g, Ajmoth Oken. (Hindi-Kusum), Butea monosperma (Lam.) Kuntze (Hindi-palas) and Zizyphus mauritiana Lam. (Hindi-Ber) are important ones [19]. The aim of (Tukhm-e-Karafs-Apium graveolens Linn., seeds) 10g, Jatamansi (Sumbul- this paper is to identify the chemical composition of the essential oil from ut-Teeb - Nardostachys jatamansi DC, rhizome) 10g, Red rose (Gul-e-Surkh- Safoof-e-Muhazzil by analysing GLC and GC-MS data for the first time. Rosa damascena Mill, petals) 25 g, Oregano (Marzanjosh- Origanum vulgare Linn., whole plant) 25 g and Lakh Maghsool (Natural resin-animal origin Laccifer lacca Kerr.)10 g. [1] Ajowan is the dried ripe seed of Trachysperum MATERIAL AND METHODS ammi (L), Sprague, belonging to the family Apiaceae. [2] It has an agreeable taste and an aromatic odour. [3] It is useful as antiseptic, antispasmodic, Collections of Plant Material and Authentication stimulant, tonic and carminative and to treat sore throat, bronchitis, diar- All the drugs were purchased from Samsi Dawakhana, Ballimaran, Delhi and rhoea and cholera. [2] Thymol, the major phenolic compound present in authenticated by Dr. H.B. Singh, Scientist F and Head, Raw Materials Her- Ajwain, has been reported to be a germicide, antispasmodic, and antifungal barium and Museum, National Institute of Science Communication and In- agent. [4] Apium graveolens L. or dill, is commonly known as Celery belong- formation Resources (NISCAIR), New Delhi. Voucher specimen of drug was ing to Apiaceae family, is an annual aromatic herb known for culinary and deposited in the Raw Materials Herbarium and Museum, National Institute medicinal use since ancient times. [5,6] In India Celery plant is grown in of Science Communication and Information Resources (NISCAIR), New Punjab, Himachal Pradesh and Uttarakhand in foot hills of Himalayas. [7,8] Delhi, with reference number Ref. NISCAIR/RHMD/consult/-2010-11/1705/ Nardostachys jatamansi DC. is a small, perennial, dwarf, hairy, rhizoma- 05. tous, herbaceous, endangered and most primitive species within family Valerianaceae. The species has very long history of use as medicine in Ayurveda, Preparation of Formulation Homeopathy, ethno-medicine and Indian System of Medicine (ISM) to modern The formulation was prepared as the method described in National Formu- medicine industry which is distributed in the Himalayas from Pakistan, India lary of Unani Medicines [1]: (Jammu and Kashmir, Himachal Pradesh, Uttarakhand, Sikkim) to Nepal, [9] 1.Nankhwah (Tachyspermum ammi L.) Seeds 10 g. Tibet and China between 3300 to 5000 m asl. Rosa damascena Mill 2.Ajmoth (Apium graveolens) Seeds 10 g. belongs to the family of Rosaceae, many species and variety of which are 3.Jatamansi (Nardostachys jatamansi) Roots 10 g. [10,11] cultivated throughout the world as an ornamental plant. Marzanjosh 4.Luk Maghsool (Purified Laccifer lacca) Resin 10 g. (Origanum vulgare L.) is an aromatic plant with a wide distribution through- 5.Gul-e-Surkh (Rosa damascena) Petals 25 g out the Mediterranean area and Asia. [12] Major components are carvacrol and 6.Marzanjosh (Origanum vulgare) Whole plant 25 g thymol that constitute about 78 to 82% of the total oil. [13] It has been suggested that the essential oil derived from O. vulgare possess in vitro Isolation of volatile oil antimicrobial [14,15], antifungal [16], insecticidal [17] and antioxidant [18] proper- The freshly prepared formulation (100 gm) was hydro-distilled for six hours with Clevenger apparatus. The yield of volatile oil obtained was 1.3 % v/w. The reddish brown coloured volatile oil was collected in the gradu- *Corresponding author. ated tube. The collected volatile oil was dried over anhydrous sodium sul- phate and stored at 4 ºC in the dark. Prof. S. H. Ansari, Department of Pharmacognosy and Phytochemistry, GC Analysis Faculty of Pharmacy, Jamia Hamdard, The gas chromatographic analysis of the volatile oil was carried out on New Delhi - 110 062, India. Shimadzu 2010 Gas Chromatograph (Japan) equipped with a flame ioniza- tion detector (FID) and AB-Innowax 7031428 WCOT fused capillary column (60 m x 0.25 mm x 0.25 μm). The injector and detector (FID)

Journal of Pharmacy Research Vol.5 Issue 1.January 2012 12-15 Kamran Javed Naquvi et al. / Journal of Pharmacy Research 2012,5(1),12-15 temperatures were maintain at 250 and 270 ºC, respectively. The carrier gas GC-MS Analysis used was nitrogen at a flow rate of 1.21 mL/min with column pressure of The analysis of the volatile constituents were run on a Shimadzu QP-2010 155.1 kPa. The sample (0.2 μl) was injected into the column with a split GC-MS system equipped with AB-Innowax 7031428 WCOT column (60 ratio of 80:1. Component separation was achieved following a linear tem- m x 0.25 mm x 0.25 μm) directly coupled to the MS. The carrier gas was perature programmed from 60-230 ºC at a rate of 3 ºC/min and then held at helium with a flow rate of 1.21 mL/min. oven temperature was programmed 230 ºC for 9 min, with a total run time of 55.14 min. Percentage of the as 50 ºC for 1 min and subsequently held isothermal for 2 min. injector port: constituents were calculated by electronic integration of FID peak areas. 250 ºC, detector: 280 ºC, split ratio 1:50, volume injected: 1 μL of the oil. The recording was performed at 70 eV, scan time 1.5 s; mass range 40-750 Table 1: Constituents of volatile oil of Safoof-e-muhazzil amu. Software adopted to handle mass spectra and chromatograph was a S. no Constituents Retention Percentage Identification Chem station. Index Identification 1. Camphene 939 0.16 KI, MS The individual peaks/constituents were identified by gas chromatography 2. L-Limonene 1014 1.39 KI, MS 3. cis-β-Ocimene 1016 0.99 KI, MS by comparison of their retention indices (R.I.) either with those of authen- 4. o-Cymene 1016 0.99 KI, MS tic compounds available in author’s laboratory or with those of literature in 5. Isoborneol 1129 0.22 KI, MS close agreement to R.I. [20-26] Further identification was made by compari- 6. t-Verbenal 1131 0.59 KI, MS son of fragmentation pattern of mass spectra obtained by GC-MS analysis 7. 4-Terpineol 1149 0.18 KI, MS 8. trans-Caryophyllene 1404 0.18 KI, MS with those stored in the spectrometer database of NBS 54 K.L, WILEY8 [20-26] 9. β –Gurjunene 1413 1.28 KI, MS libraries and published literature. Retention indices of the components 10. Germacrenol D 1458 0.22 KI, MS were determined relative to the retention times of a series of n-alkanes 11. β –Selinene 1464 0.21 KI, MS relative to C -C on HPS and HP-20M columns. 12. -Guaiene 1480 0.25 KI, MS 9 20 13. Verbenone 1481 0.10 KI, MS 14. Germacrene D 1483 0.23 KI, MS RESULTS AND DISCUSSION 15. Aromadendrene 1495 0.06 KI, MS Volatile oil of Safoof-e-Muhazzil was characterized by less number of monot- 16. β- Besabolene 1498 9.73 KI, MS erpenes (11.62 %) and a large number of sesquiterpenes (85.05 %) with 17. - Cadinene 1502 0.78 KI, MS 18.  – Cadinene 1518 1.37 KI, MS bisabolene (9.73 %), eudes-4(14),11-diene (28.61 %), globulol (9.13 %), t- 19. Germacren-4-ol 1530 0.50 KI, MS cadinol (4.15 %), viridiflorol laurate (16.40 %) types. Among thirteen monot- 20. trans-Cadina-1,4-diene 1535 2.08 KI, MS erpenes (11.62 %), twelve were positively characterized and one was par- 21. Elemicin 1555 0.48 KI, MS tially characterized. There were four each monoterpene hydrocarbons (3.53 22. Eudes-4(14),11-diene 1559 28.61 KI, MS 23. β – Germacrene 1561 0.08 KI, MS %), three alcohols (6.54 %), two esters (0.86 %), one each ketone (0.10 %), 24. cis-Nerolidol 1563 0.08 KI, MS two monoterpene aldehydes (0.67 %) and one each monoterpene ether 25. -Germacren-4-ol 1576 0.52 KI, MS (0.48 %). The prominent monoterpenes were thymol (6.14 %) followed by 26. Globulol 1578 9.13 KI, MS L-limonene (1.39 %), cis-β-ocimene (0.99 %), o-ocimene (0.99 %). 27. 4-Isopropylbenzaldehyde 1579 0.08 KI, MS 28. Viridiflorol 1585 0.37 KI, MS 29. cis-Carveol pentanoate 1593 0.48 KI, MS Among forty four sesquiterpenes (85.05 %), thirty seven components 30. Viridiflorene epoxide 1598 0.70 KI, MS (84.56 %) were positively identified and seven components were partially 31. Humulene epoxide 1601 0.16 KI, MS 32. Viridiflorene epoxide 1605 0.08 KI, MS characterized. There were eighteen sesquiterpene hydrocarbons (46.67 %), 33. -Eudesmol (Junifer camphor) 1615 0.26 KI, MS twenty were sesquiterpene alcohol (20.49 %), three esters (16.77 %) and 34. Eudesmol 1630 0.12 KI, MS three sesquiterpenes ketones (1.12 %). The predominant sesquiterpenes 35. Isosphathulenol 1634 0.31 KI, MS were eudes-4(14),11-diene (28.61 %), viridiflorol laurate (16.40 %), 36. Caryophyllenol 1635 0.05 KI, MS 37. t-Muurolol 1642 0.14 KI, MS bisabolene (9.73 %), globol (9.13 %), t-cadinol (4.15 %), trans-cadine-1,4- 38. t-Cadinol 1645 4.15 KI, MS diene (2.08 %), 2E, 6E-farnesol (1.41), - cadinene (1.37 %) and β-gurjunene 39. Bisabolol oxide 1658 0.37 KI, MS (1.28). The partially identified sesquiterpenes included β-germacrene, cis- 40. Caryophylla-4,8-diene-5-ol 1661 0.15 KI, MS nerolidol, viridiflorene epoxide, caryophyllenol, cubenol, 4-methyl-4-phe 41. Valeranone 1668 0.73 KI, MS 42. Patchouli alcohol 1668 0.35 KI, MS nyl-2,3:5,6-diepoxycyclohexanone and all of them occurred in a very less 43. Aromadendrene oxide 1669 0.19 KI, MS amount (< 0.1%). The sesquiterpenes present in trace amount were trans- 44. Ledene oxide II 1670 0.46 KI, MS caryophyllene, β-gurjunene, germacrenol D, β-selinene, δ-guaiene, verbenone, 45. Cubenol 1670 0.07 KI, MS 46. Salina-6-en-4-ol 1675 0.42 KI, MS germacrene D, -cadinene, -cadinene, germacren-4-ol, trans-cadina-1,4-di- 47. -Bisabolol 1669 0.93 KI, MS ene, eudes-4(14),11-diene, cis-nerolidol, -germacren-4-ol, viridiflorol, cis- 48. Thymol 1675 6.14 KI, MS carveol pentanoate, viridiflorene epoxide, humulene epoxide, viridiflorene 49. (E,E)-Germacra-3,7(11),9-trien-6-one 1679 0.24 KI, MS epoxide, -eudesmol (junifer camphor), eudesmol, isosphathulenol, t- 50. Thymol stearate 1701 0.38 KI, MS 51. (2 Z, 6 E)-Farnesol 1722 0.68 KI, MS muurolol, bisabolol oxide, caryophylla-4,8-diene-5-ol, valeranone, patchouli 52. (2E, 6E)-Farnesol 1733 1.41 KI, MS alcohol, aromadendrene oxide, ledene oxide II, salina-6-en-4-ol, α-bisabolol, 53. -Turmenone 1745 0.15 KI, MS (E,E)-germacra-3,7(11),9-trien-6-one, (2 Z, 6 E)-farnesol, (2E, 6E)-farnesol, 54. n-Heptadecane 1780 0.16 KI, MS 55. Neoclovenoxide alcohol 1810 0.24 KI, MS α-turmenone, n-heptadecane, neoclovenoxide alcohol, 4-methyl-4-phenyl- 56. 4-Methyl-4-phenyl-2,3:5,6- 1821 0.06 KI, MS 2,3:5,6-diepoxycyclohexanone, valerenol benzoate, veridiflorol glycolate. diepoxycyclohexanone Only two diterpene alcohol (0.56 %), trans-phytol and 13(16), 14-labdiene- 57. Valerenol benzoate 1835 0.16 KI, MS 58. n-Eicos-15-enol 1846 0.47 KI, MS 8-ol were identified and both of them occurred in trace amount. n-eicos-15- 59. 2-Benzlidine 1855 0.09 KI, MS enol, 9,12-octadecadienoic acid methyl ester, n-heptacosane and methyl-6- 60. Viridiflorol laurate 1925 16.40 KI, MS 61. 9,12-Octadecadienoic acid methyl ester 1966 0.17 KI, MS hydroxyoctadecanoate were the aliphatic components identified and 2- 62. n-Heptacosane 2001 0.14 KI, MS benzilidine and amyrolin were the aromatic components identified. Among 63. Methyl-6-hydroxy-octadecanoate 2005 0.19 KI, MS sixty seven volatile oil constituents, fifty eight components (99.35 %) were 64. trans-Phytol 2011 0.15 KI, MS 65. 13(16), 14-Labdien-8-ol 2035 0.41 KI, MS completely identified and nine volatile oil constituents (0.65 %) comprising 66. Veridiflorol glycolate 2115 0.21 KI, MS were partially characterized in the sample. 67. Amyrolin 2160 0.52 KI, MS Journal of Pharmacy Research Vol.5 Issue 1.January 2012 12-15 Kamran Javed Naquvi et al. / Journal of Pharmacy Research 2012,5(1),12-15

Fig. 1. GC-MS Spectra of Safoof-e-Muhazzil Fig. 2. GC-Spectra of Safoof-e-Muhazzil

H H H H

Limonene cis-β-Ocimene o-cymene β-Gurjurene β-Bisabolene -Cadinene

O H

H HO

trans-Cadina-1,4-diene Eudesma-4(14),11-diene Globulol t-Cadinol Thymol

O H

H O

2E, 6E-Farnesol Viridiflorol Aromadendrene Salina-6-en-4-ol

Fig. 3. Prominent components of volatile oil Safoof-e-muhazzil

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Journal of Pharmacy Research Vol.5 Issue 1.January 2012 12-15