SUPPLEMENTARY MATERIAL
Identification of chemical compounds present in different fractions of Annona reticulata L.
leaf by using GC- MS
Soumya P. Rout 1 *, Durga M. Kar 1
1
Siksha 'O' Anusandhan University, Department of Pharmacology, School of Pharmaceutical
Sciences, Kalinga Nagar, Ghatikia, Bhubaneswar-751003, Odisha, India
* Corresponding Author
Email: spr_cology@rediffmail.com
Abstract
Leave of Annona reticulata Linn (Family - Annonaceae), is used as suppurative, stimulant,
antispasmodic, sudorific, anthelmintic, insecticidal in ethno-medicinal system. The leave extracts
also has been shown to have antidiabetic and antioxidant property. The present investigation had
been carried out to identify presence of different chemical compounds in several fractions
prepared from
Hydro-Alcoholic extract
of
Annona reticulata leaf by using Gas
Chromatography–Mass Spectrometry Technique. The mass spectra of the compounds found in
the fractions were matched with the National Institute of Standards and Technology (NIST)
library. GC-MS analysis of fractions prepared from Hydro-Alcoholic extract of Annona
reticulata leaf revealed presence of: 9, 10-Dimethyltricyclo [4.2.1.1(2, 5)] decane-9, 10-diol and
4-(1,5-Dihydroxy-2,6,6-trimethylcyclohex-2-enyl)but-3-en-2-one
as
major
compounds
in
Chloroform Fraction; 3,7-Dimethyl-6-nonen-1-ol acetate and 9-Octadecenamide,(Z)- as major
compounds in Ethylacetate Fraction; Glycerin and D-Glucose, 6-O-α-D-galactopyranosyl- as
major compounds in Methanol Fraction; Desulphosinigrin and α -Methyl-D-mannopyranoside as
major compounds in the residual or Hydro-Alcoholic fraction.
Keywords: Annona reticulata, GC-MS technique, Antidiabetic
Experimental
Plant collection, Preparation of Extract and its Fractions: The aerial parts of A. reticulata L.
were collected in the month of July – August 2010, from the rural area of the dist. Cuttack,
Odisha, India and authenticated by Dr. Kshetra Mohan Das, Central Rice Research Institute,
Cuttack, Odisha, India and a specimen voucher (Specimen No. 10-11/SPS/SOAU) has been kept
in the University for future reference. The leaves were shade dried, made into coarse powder and
the powdered material was initially defatted with petroleum ether and then subjected to cold
maceration process for 72-hours by using 1: 1 mixture of methanol and water as solvent to
prepare hydoalcoholic extract of Annona reticlulata leave. The extract was filtered and
concentrated by rotary evaporator. For preparation of different fractions, method published by
Sourav et al., was used.The extract is mixed with water and methanol (1:1) and used for
fractionation by using chloroform, ethylacetate and methanol as solvents by using separating
funnel (Sourav
et al., 2012). The different fractions prepared are; Chloroform fraction of
hydroalcoholic extract of Annona reticulata, Ethylacetae fraction of hydroalcoholic extract of
Annona reticulata, Methanolic fraction of hydroalcoholic extract of Annona reticulata. The
residue is also taken for analysis and named as Hydro-Alcoholic fraction of hydroalcoholic
extract of Annona reticulata.
GC-MS Analysis
GC-MS analysis was done by using GC Clarus 500 Perkin Elmer system and gas chromatograph
interfaced with mass detector Turbo mass gold-Perkin Elmer (GC-MS) (Hema et al., 2010). The
following programming condition has been employed;
GC Programme - Column: Elite-5MS (5% Diphenyl / 95% Dimethyl poly siloxane), 30 x
0.25mm x 0.25m df, Carrier gas: Helium (99.999%) with constant flow rate of 1ml per min,
(Split ratio: 10:1), Sample Injection volume 2μl, Software: Turbomass 5.2,Oven operating in
electron impact mode at 70 eV, oven temperature was fixed from 110°C (isothermal for 2 min.),
with an increase rate of 10°C/min upto 200°C with no hold and then at rate of 5°C/min upto
280°C ending with a 9 min hold. Injector temperature was 2500C, Ion-source temperature 280°C
and total GC running time was 36minutes.
MS Programme - Library used NIST Version-Year 2005, Inlet line temperature 200° C, Source
temperature 200 ° C, Electron energy: 70 eV, Mass scan (m/z): 45-450, Solvent Delay: 0-2 min,
Total MS running time: 36 min.
Identification of Compounds
Interpretation on mass spectrum GC-MS was done by using National Institute Standard and
Technology (NIST) database 2005. The spectra of unknown compounds were compared with
that of known compounds stored in the NIST library. The Name, Molecular weight and Structure
of the components of the test materials were confirmed (Hema et al., 2010). Presence of twentyfive compounds in chloroform fraction, eighteen compounds in ethylacetate fraction, ten
compounds in methanol fraction and fifteen compounds in hydro-alcoholic fraction have been
identified.
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