INTRODUCTION:

Bombax ceiba Linn.(Syn. B. Malabaricum Dc.) Merr. and Gossamprinus malabaric (Dc) Merr., (Fam: Bombacaceae) is lofty deciduous, buttressed tree with deep red flowers¹. Young Leaves and Flower buds are edible². Extensive reports are available on the phytochemical and pharmacognostic characters of the plant.^3,4.

MATERIALS AND METHODS:

Extraction and Fractionation:

The gynaceum part of B.ceiba has been chosen for the investigation purpose. The gynaceum part of the fresh flowers of B.ceiba has been collected in and around Kumbakonam, Tamil Nadu. The gynaceum were extracted with 90% MeOH under reflux. The MeOH extract was concentrated in vacuo and fractionated with the solvents like benzene, peroxide free Et₂O and EtOAc.

Et₂O Fraction: ( Flavonol – Quercetagetin ):

The Et₂O Fraction was concentrated in vacuo and kept in an ice chest for about a week. A yellow solid that separated was filtered and examined. It came out as yellow crystals. On recrystalization from MeOH, it had λ_max nm 260,290sh, 3 65;+NaOMe 268,325,345sh, 420;+AlCl₃ 270,300sh, 360sh, 420;+( AlCl₃ + HCl ) 270, 300sh, 60sh, 370sh, 400;+NaOAc270, 300sh, 335sh,400;+( NaOAc+H₃BO₃) 265,300sh,370. It was identified as Quercetagetin, by comparing it with an authentic sample isolated from the flowers of Tagetes erecta⁵.

It gave an orange yellow precipitate with basic Lead-Acetate with diliute alkali it gave yellow solutions that rapidly changed to green. It responded to Horhammer-Hansel⁶, Willson’s boric acid⁷ and Gibb’s⁸ tests. It did not respond to Molisch’s test. It appeared yellow under UV with or without NH_3. It had R_fvalues depicted in table1

EtOAc Fraction: - (Quercetagetin -3-O-D-

glucofuranoside):

The EtOAc fraction was concentrated in vacuo and left in an ice-chest for a few days. A yellow solid that separated was filtered and studied. Pale yellow leaflets came out with M.P.203⁰C on recrystallisation from MeOH. It was soluble in EtOH and EtOAc but insoluble in cold water. It developed a green color with alc. Fe³⁺. Pink colour with Mg-HCl and a yellow precipitate with aq.lead acetate. It reduced Amm.AgNO₃solution but not Fehiling’s solution. It appeared deep purple under UV that turned yellowish green on exposure to NH₃. It reponded to Wilson’s boric acid, Molisch’s and Gibb’s tests but did not answer the Horhammer-Hansel test. It had λ_max nm 261,270sh, 3 55;+NaOMe 273,338, 410;+AlCl₃ 278,302sh, 339sh, 427;+( AlCl₃ + HCl ) 269, 281sh, 301sh, 373, 407sh;+NaOAc 273, 326sh, 385sh;+( NaOAc+H₃BO₃) 265,365.It had R_f values depicted in table1

Hydrolysis of the glycoside:-

The glycoside dissolved in hot aq.MeOH was hydrolysed with H₂SO₄ at 100⁰C for about 2 hr and the hydrolytic products identified as described below.

Table – 1: Rf ( X 100 ) values of the constituents of the flowers of B. Ceiba ( Whatman No. 1, Ascending 30 ± 2⁰ c )

Compound	a	b	c	d	e	f	g	h	i
Aglycone from Et₂O fraction	01	04	10	16	30	57	31	35	42
Quercetagetin ( authentic )	01	04	10	16	30	57	31	35	42
Glycoside from EtOAC fraction	18	05	20	30	60	75	80	76	40

Solvent Key:-

a) H₂O b) 5% aq.HOAc, c)15% aq.HOAc, d)30% aq. HOAc,

e) 60%aq.HOAc, f) BAW ( n-BuOH:HOAc:H₂O=4:1:5 ) g) water saturated phenol

h) t-BuOH:HOAc:H₂O=3:1:1 i) Forestal ( HOAc:con.HCl:H2O=30:3:10)

Table – 2 ¹³C-NMR spectral data and their assignment for the glycoside from the flowers of B.Ceiba

Carbon Number	Quercetagetin ( from literature δ ppm )	Glycoside isolated from EtOAc fraction (δ ppm)
C2	156.3	156.4
C3	133.4	133.8
C4	177.4	175.5
C5	161.2	161.1
C6	120.3	120.0
C7	164.1	166.1
C8	115.2	115.1
C9	156.3	156.4
C10	115.1	115.1
C1’	121.6	121.6
C2’	115.2	115.1
C3’	144.8	144.6
C4’	148.4	149.8
C5’	116.2	116.5
C6’	121.2	121.6
C1’’	110.0	113.8
C2’’	74.0	73.7
C3’’	76.4	76.3
C4’’	69.9	69.7
C5’’	77.3	77.3
C6’’	60.9	60.5

Identification of Aglycone :- ( Flavonol-Quercetagetin):

The aglycone on recrystallisation from MeOH afforded a yellow crystalline solid, M.P.324-325⁰C which was identified as quercetagetin by colour reaction, behaviour under UV and R_f. It had the same UV spectral values, mentioned under Et₂O fraction.

Identification of Sugar :- ( glucose):

The aq.hydrolysite after the removal of the aglycone was neutralized with BaCO₃ and filtered. The concentrated filterate on PC gave R_f values corresponding to those of glucose. The glycoside was thus identified as quercetagetin -3-O-D-glucofuranoside and this was confirmed by co-PC with an authentic sample of Teges erecta.

RESULTS AND DISCUSSION:

The Gynaceum parts of the fresh flowers of Bombax ceiba have been found to contain quercetagetin and its glycoside quercetagetin -3-O-D-glucofuranoside.

The UV spectrum of the Et₂O fraction showed two major absorption peaks at 365nm and 260nm showing a flavonol skeleton. A bathochromic shift of 55nm in band I observed in its NaOMe spectrum indicated the presence of a free – 4’ OH group. The AlCl₃ / HCl spectra of the aglycone showed

five absorption peaks indicating a free -5 OH group. It exhibited a fluorescent light blue under UV with or without NH₃. A bathochromic shift of +35 nm in AlCl₃/HCl spectra was an evidence for a free 5-OH in aglycone. The presence of free C-7 OH is evident from a shift of +10 nm on the addition of NaOAc. The presence of a o-dihydroxyl group in both A and B rings could be infered from a shift of +5 nm on the addition of H₃BO₃. In the AlCl₃ spectrum an absorption peak was noticed at 420nm which an addition of HCl reduced by 20nm. This is another evidence for the presence of catechol type dihydroxyl group in the A as well as B rings.

The UV spectrum of the glycoside exhibited two major absorption peaks at 355nm and 261nm. A comparison of band I absorption of the glycoside and that of the aglycone revealed that there may be 3-glycosylation in the flavonol. A bathochromic shift of +55nm in NaOMe confirmed the presence of a free –OH at C 4’. The AlCl₃ spectra showed 4 absorption peaks to reveal the presence of a free 5-OH group. It was conformed by the bathochromic shift of +52nm on the addition of AlCl₃/HCl in the glycoside. The presence of a free –OH at C7 was evident from the +12 nm shift on the addition of NaOAc. The band I absorption in AlCl₃ spectrum is +20nm more than that noticed on the addition of AlCl₃/HCl. This is indicative of the existence of o-dihydroxyl groups in A and B rings.

In the ¹H NMR spectrum. The A ring protons at C6 and C8 appear at δ 6.22 and 6.45ppm respectively. The 5-OH protons resonate at δ 12.55ppm. The proton at C 5’ appears at δ 6.80ppm. The protons at C 2’ and C 6’ appear at δ 7.29 and 7.59ppm. The H 1” of the glucoside resonates at δ 5.20ppm. The remaining sugar protons appear in the range of 3.28 to 3.39ppm.

Supporting evidence for the structure of the glycoside was provided by the analysis of of ¹³C NMR (100 MHz, DMSO-d₆, TMS) data and a complete assignment is given in table 2.

Based on this the aglycone and the glycoside have been characterized as quercetagetin and quercetagetin -3-O-D-glucofuranoside. The structure showed in the figure 1.

CONCLUSION:

Bombax ceiba is a buttressed tree with deep red flowers. The gynaceum part of the flowers was taken for analysis. on phytochemical investigation of the EtOAc soluble fraction yielded quercetagetin -3-O-β-D-glucofuranoside.

ACKNOWLEDGEMENTS:

The authors wish to acknowledge Indian Institute of Science, Bangalore and National Institute of Technology, Trichirappalli for technical assistance.

REFERENCES:

1. J.D.Hooker, The Flora of British India,L.Reeve and Co.London, 1954,1,349

2. The Wealth of India, Raw materials, CSIR , New Delhi, 1972, 9, 177

3. P.N.Mehra and C.R.Karnik, Indian J Pharm., 1968, 30,284

4. H.N.R. Chaudhuri, Bull Bot.Soc.Bengal, 1971, 25, 37\

5. P.S.Rao and T.R.Seshadri Proc.Indian Acad.sci., 1941, 14A, 289

6. L.Horammer and R.Hansel, Arch.Pharm.Berl., 1955, 288, 315

7. C.W. Wilson, J.Amer. Chem. Soc., 1939, 61, 2303

8. F.E.King, T.J.King and L.C.Manning J.Chem.Soc., 1957,563

Received on 11.09.2009 Modified on 09.11.2009

Asian J. Research Chem. 3(1): Jan.-Mar. 2010; Page 78-80