Phytochemical screening and GC MS analysis of methanolic extract of Abelmoschus moschatus

The study was designed to evaluate the phytoconstituents present in the methanolic extract of aerial parts of Abelmoschus moschatus. The qualitative phytochemical screening of different extracts of aerial parts revealed the presence of somebioactive compounds. GC –MSanalysiswasperformedusing Shimadzu Gas chromatography-mass spectroscopy (Model Number: QP2010S) instrument. GC-MS detection of phytoconstituents was done by computer evaluation ofmass spectra of samples throughNational Institute Standard and Technology (NIST II) andWILEY 8 library. GC –MS analysis detected the presence of 14 compounds. GC – MS pro ile of the methanolic extract revealed the presence ofmegastigmatrienone, phytol, loliolide, farnesyl acetate, methyl linoleates, gamma-sitosterol, cis, cis, cis-7,10,13-Hexadecatrienal, thymine, pyranone, coumarin, 2 – methoxy 4 – vinyl phenol, guanosine, chinasaure and 3cyclopentyl propionic acid 2 dimethyl aminoethyl ester. The current study suggests that methanolic extracts of aerial parts of Abelmoschus moschatus contain phytoconstituents with antioxidant and cytoprotective activity. The study results will pave a way for the production of therapeutic agents which can be used for the treatment of various diseases.


INTRODUCTION
The genus Abelmoschus comprise of about 15 species of lowering plants and belongs to Malvaceae family. It is an aromatic and medicinal oil yielding plant commonly grown in some parts of our country. (Pawar and Vyawahare, 2018). Abel-moschus moschatus has lots of medicinal properties and is called as Kattukasthuri or musk mallow in different areas. The plant is widely used in folk medicine for its therapeutic properties. Almost every part of musk mallow has medicinal properties. The plant has a long record of use in conventional health care systems. Many races think this plant as a remedy for various diseases. In India, roots, leaves, and seeds of this plant are used for therapeutic purpose . Mucilage prepared from root and leaves of the plant is advocated in sexually transmitted bacterial infections. The poultice of leaves and roots is used in cystitis, fever, headache, rheumatism, and for varicose veins and boils. In Egypt, seeds are munched to alleviate gastric disorders. Different types of ayurvedic formulations from seeds are used for nervous weakness, hysteria and other neurological conditions. Traditionally seeds have considerable role in the therapy of progressive loss of neurological functions. Seeds rubbed to a paste with milk are used to cure itch (Padua et al., 1999).

Figure 1: Abelmoschus moschatus
Flower infusion is contraceptive. The mucilaginous seeds are emollient and demulcent. . Topical application of the seed oil relieves abdominal cramps, circulatory insuf iciency and joint pain. Ambrette oil has insecticidal and sexual stimulant properties (Dwivedi and Argal, 2015;Purohit and Vyas, 2004). Although the plant has enormous use in traditional medicine, most of the biological activities are not scienti ically validated. Abelmoschus moschatus is a perennial plant. Stems are solid and is about 30cm tall. Leaves have variable size and shape and some are sagittate. Leaf margins are lobed. Leaf apex is obtuse. Flowers have a diameter of 4 -5cm. They are solitary. The lowers have 5 petals with colours ranging from white, pale yellow to dark pink Figure 1. They last for only one day and their lowering depends on the timing of the wet season. Seeds are held in hairy tough, papery capsules. The seeds have a sweet, lowery, heavy fragrance similar to that of musk (Harborne, 1998)

Collection of the plant
The plant Abelmoschus moschatus Figure 1 was collected from Pala, Kottayam district, Kerala. The plant was authenticated at Botanical Survey of India, and deposited in the herbarium with voucher No. BSI/SRC/5/23/2020/Tech/65

Chemicals and Reagents
All the chemicals and reagents were procured from certi ied suppliers and of analytical reagent grade.

Preparation of the extracts
The plant extracts were prepared following the procedure of Gopalasatheeskumar (2018) by Soxhlet extraction method. The aerial parts of the plant were dried under shade and was ground to a powder using an electrical blender. Extraction was carried out by continuous hot percolation method by using the following solvents in order i.e., Petroleum ether, Chloroform, Acetone and Methanol. The extracts were then concentrated using a rotary evaporator and kept at 4 o C until used. These extracts were used for phytochemical screening.

Phytochemical screening
Phytochemical analysis of crude extracts of aerial parts of Abelmoschus moschatus was carried out as per standard procedures (Evans and Trease, 2009;Sofowora, 1993).

GC MS analysis
GC -MS analysis was done on the methanolic extract of aerial parts of Abelmoschus moschatus. Shimadzu GC -MS (Model Number: QP2010S) instrument with GC -MS solutions software was used for analysis. The oven temperature is maintained at 280 o C at a rate of 5 o c/min. Elite -5MS column of 30m length,0.25mmID and 0.25micrometer thickness was used.

Figure 2: GC -MS chromatogram
Detection of the phytoconstituents present in the methanolic extract of Abelmoschus moschatus was based on the comparison of mass spectra of samples through National Institute Standard and Technology (NIST II) and WILEY 8 library. Parameters such as comparison of peaks and retention time, computer matching and the characteristic fragmentation patterns of the mass spectra were used for characterizing the phytoconstituents. GC -MS pro ile of the compounds identi ied is given in Figure 2.  1  Alkaloids  ----2  Glycosides  ----3 Steroids and triterpenoids + -+ + 4 Flavanoids -+ -+ 5 Carbohydrates + + + + 6 Phenolic compounds and tannins -- Fixed oils and fats -----Absent, + Present, PEE -Petroleum Ether Extract, CE-Chloroform Extract, AE -Acetone Extract, ME -Methanol Extract

RESULTS AND DISCUSSION
The phytoconstituents present in the extracts were qualitatively analyzed using chemical tests. Phytochemical analysis of the four extracts of Abelmoschus moschatus showed the presence of steroids, Triterpenoids, Flavonoids and carbohydrates Table 1.
The methanolic extract contains maximum number of phytoconstituents and GC -MS technique was used to identify the constituents in this extract. The results of GC -MS analysis is shown in Table 2.
Peak area and retention time were used for the identi ication of compounds. GC-MS analysis of methanolic extract of aerial parts of abelmoschus moschatus revealed the presence of terpenes (Megastigmatrienone, Phytol).
Megastigmatrienones are nor -isoprenoid compounds derived from carotenoids and are the main compounds found in tobacco. (Anthony et al., 2009) Phytol is an acyclic diterpene alcohol. Phytol has been reported to have antioxidant, anti-in lammatory and antitumor effects. Hydroxyl group present in phytol confers antioxidant property to the molecule (Guimarães et al., 2010). Loliolide is a monoterpene lactone and possess signi icant antioxidant and cell protective effects against the cellular damage produced by H 2 O 2 . (Yang et al., 2011). Gamma sitosterol is an important plant sterol. Gamma-sitosterol exhibited hypoglycemic effect on streptozocin induced diabetes in rats. Gamma sitosterol is reported to increase insulin secretion and inhibit glucogenesis.
Literature survey reveals that gamma sitosterol has a role in the activation of components involved in extrinsic apoptotic pathway in human lung and breast ade-nocarcinoma cells. (Sundarraj et al., 2012). Farnesyl acetate and methyl linoleate are fatty acid esters and possess activity against gram -ve bacteria, moulds and dermatophytes. (Pauli, 2001)

CONCLUSIONS
The presence of various bioactive constituents like terpenes, monoterpene lactone, steroids and carotenoid derivatives in Abelmoschus moschatus validates the use of this medicinal herb for a number of diseases by traditional healers. However, isolation of individual phytochemical constituents and screening for their bioactivity will give hopeful results. Therefore, Abelmoschus moschatus can be considered as a plant of considerable phytopharmaceutical importance.