Analysis of Spectral Data of the Chemical Constituents from the Leaves of Jasminum grandiflorum L . , Achyranthes aspera L . and Tinospora cordifolia ( Willd . ) Miers

Jasminum grandiflorum L. (Oleaceae) is used to relieve coughs, depression, dizziness, eye diseases, facial paralysis, general debility, fevers, headache, psoriasis, sciatica, skin diseases and vertigo. Achyranthes aspera L. (Amaranthaceae) is utilized to treat anorexia, ascites, respiratory problems, kidney, brain and skin diseases, cholera, convulsions, diabetes, fistula, hysteria, insect bites, malaria, night blindness, obesity, piles, snake bites, stomach disorders, swellings, tooth aches, tumors and wounds. Tinospora cordifolia (Willd.) Miers (Menispermaceae) is effective to alleviate anemia, debility, diabetes, diarrhea, dysentery, dyspepsia, fevers, jaundice, rheumatism, urinary and skin diseases, scorpion stings and snake bites. The air-dried plant leaves were exhaustively extracted with methanol individually in a Soxhlet apparatus. The concentrated methanol extracts were adsorbed on silica gel for column and chromatographed over silica gel column separately. The columns were eluted with petroleum ether, chloroform and methanol successively to isolate the phytoconstituents. Phytochemical investigation of the leaves of J. grandiflorum afforded glyceryl behenate (2,3-dihydroxypropyl docosanoate, 1), glycerol cerotate (2,3-dihydroxypropyl 1-hexacosanoate, 2), cerotyl O-β-D-diarabinoside (nhexacosanoyl-O-β-Darabinopyranosyl-(2ʹ→1′′)-O-β-Darabinopyranoside / cerotyl O-β-D-arabinopyranosyl-(2ʹ→1ʹʹ)-O-β-D-arabinopyranoside, 3), stearyl-O-α-Dtriglucoside (stearyl glucopyranosyl-(6′→1′′)-O-α-Dglucopyranosyl-(6′′→1′′′)-O-α-Dglucopyranoside, 4) and behenyl-O-α-D-glucopyranosyl-(6’→1′′)-O-α-Dglucopyranosyl-(6’′→1′′′)-O-α-D-glucopyranoside, 5). The leaves of A. aspera and T. cordifolia furnished a new diterpenoid ester aromadendr-10(14)-en-15-olyl (E)-ferulate 6) and an aromatic ester phenyl ethyl behenate 7), respectively. The structures of these phytoconstituents have been established on the basis of spectral data analysis and glycosidic and phenolic chemical reactions.


General Procedures
Melting points were determined on a Perfit melting point apparatus and are uncorrected.UV spectra were determined on Shimadzu-120 double beam spectrophotometer with methanol as a solvent.IR spectra were recorded in KBr pellet on a Shimadzu FTIR-8400 spectrophotometer.The 1 H NMR (300 MHz) and 13 C NMR (75 MHz) spectra were scanned on a Bruker DRX instruments using TMS as an internal standard and coupling constants (J values) are expressed in Hertz (Hz).Mass spectra were recorded by affecting electron impact ionization at 70 eV on a Jeol SX-102 mass spectrometer equipped with direct inlet prob system.The m/z values of the more intense peaks are mentioned and the figures in bracket attached to each m/z values indicated relative intensities with respect to the base peak.Column chromatography was performed on silica gel (60-120 mesh; Qualigen, Mumbai, India).TLC was run on silica gel G 60 F254 precoated TLC plates (Merck, Mumbai, India).Spots were visualised by exposing to iodine vapours and UV radiations (254 and 366 nm) and spraying with ceric sulphate solution.

Plant Material
The leaves of J. grandiflorum, A. aspera and T. cordifolia, were collected locally from Delhi and authenticated by Prof. M. P. Sharma, Taxonomist, Department of Botany, Jamia Hamdard, New Delhi.The voucher specimens of these plant parts are preserved in the herbarium of the Department of Pharmacognosy and Phytochemistry, Jamia Hamdard, New Delhi.

Extraction and Isolation
The leaves of each plant (1 kg) were coarsely powdered and extracted exhaustively with methanol individually in a Soxhlet apparatus.The extracts were concentrated under reduced pressure to get dark brown masses, 110.3 g, 129.4 g and 119.7 g, respectively.The dried residue (100 g each) was dissolved in a minimum amount of methanol and adsorbed on silica gel column grade (60-120 mesh) individually to obtain a slurry.Each slurry was air-dried and chromatographed over silica gel columns loaded in petroleum ether (b.p. 60 -80°C) separately.The columns were eluted with petroleum ether, petroleum ether -chloroform (9:1, 3:1, 1:1, 1:3, v/v), chloroform and chloroform -methanol (99:1, 49:1, 19:5, 9:1, 17:3, 4:1 7:3, 1:1, v/v) mixtures.Various fractions were collected singly and matched by TLC to check homogeneity.Similar fractions having the same Rf values were combined and crystallized with solvents.The isolated compounds were recrystallized to get the following pure compounds:

Glycerol cerotate (2)
Further elution of the column with ethyl acetate -methanol
Compound 2, designated as glycerol cerotate, showed IR absorption bands for hydroxyl groups (3415, 3355 cm - 1 ), ester group (1721 cm -1 ) and long aliphatic chain (720 cm -1 ).On the basis of mass and 13 C NMR spectra its molecular ion peak was determined at m/z 470 corresponding to a molecular formula of an acyl glycerol C29H58O4.An ion peak arising at m/z 379 [C1′ -O fission, CH3-(CH2)24-CO] + indicated that cerotic acid was linked with glycerol.The 1 H NMR spectrum of 2 exhibited a multiplet at δ 4.43 and a doublet at δ 3.44 (J = 5.6 Hz) integrating for two protons each assigned to oxymethylene H2-1 and H2-3 protons, respectively, a one-proton multiplet at δ 4.06 ascribed to hydroxymethine H-2 proton, a three-proton triplet at δ 0.82 (J = 6.5 Hz) attributed to terminal C-26′ primary methyl protons and the methylene protons in the range from δ 2.51 to 1.21.The 13   suggested that the attachment of cerotic acid at C-1.The absence of any signal beyond δ 4.43 in the 1 H NMR spectrum and between δ 170.11 -71.23 in the 13 C NMR spectrum supported saturated nature of the molecule.On the basis of these evidences the structure of 2 has been elucidated 2,3-dihydroxypropyl 1-hexacosanoate (Figure 1).

Figure 2 .
Chemical constituent 6 isolated from the leaves of Achyranthes aspera C NMR spectrum of 2 CH 3 (CH 2 ) 20 CO O CH 2 CHOH CH 2 OH