Indian Journal of Chemistry

Vol. 42B, June 2003, pp. 1 5 14- 1 5 1 8

Steroid and nitro phenol esters from Bignonia unguis-cati roots

b
B iswanath Dinda*, Utpal Chandra De, Basudev Acharia , Shiho Arimab , Nariko Sato b & Yoshihiro Harigaya
Department of Chemistry, Tripura University, Suryamaninagar 799 1 30, India

E.mail: dindabtu @rediffmail.com

aDepartment of Chemistry, Indian Institute of Chemical Biology, Kolkata 700 032, India
bSchool of Pharmaceutical Sciences, Kitasato University, Minato-ku, Tokyo 1 08 864 1 , Japan

Received 21 February 2002; accepted (revised) 1 January 2003

Two new esters, I)-sitosterol cerotate 1 and 2-(4-hydroxy-3-nitrophenyl)ethyl stearate 2 have been isolated from the
roots of Bignonia unguis-cati (Bignoniaceae) and their structures are elucidated by spectroscopic and chemical studies.

4
Bignonia unguis-cati L. syn. B. gracilis Lodd syn. and 4.5 Hz, H-6) suggesting an sterol ester skeleton .
Doxantha unguis Miers (Bignoniaceae), an American This was supported by the appearance of base peak
species is grown in Indian gardens for its use in tradi­ ion at mJz 396 in the ElMS resulting from the loss of
tional system of Indian medicine l . The roots are used acid residue by McLafferty cleavage. The ElMS also
in the treatment of ulcers. Earlier investigations on recorded some mass peaks at mJz 777(M-Me, 5),
23
aerial parts of the plant reported . the presence of la­ 763(M-Et, 8), 749(M-43, 1 0) , 456( 1 5), 424( 1 2),
pachol, ceryl alcohol, j3-amyrin, j3-sitosterol, octaco­ 4 1 3(8), 397( 1 00), 3 8 1 (396-Me, 1 7) , 370(34), 35 1 (4),
sanol, tectoI and ellagic acid. We i nvestigated the 342(20), 288(5), 276( 1 0) , 2 1 4(7), 148( 1 2), 60(4),
roots of this plant and isolated two new compounds - 57(42), 43(34), which were characteristic of steroid
s6
j3-sitosterol cerotate 1 and 2-( 4-hydroxy-3- fragmentation (Scheme l) . . The mass peak at mJz
nitrophenyl)ethyl stearate 2. In this paper, we wish to 424 was characteristic of ""S steroid7. The strong mass
report the i solation and structure elucidation of these peak at mJz 397 suggested the presence of cerotic acid
compounds. 3 as an acid residue. The alkaline hydrolysis of 1 with
I N methanolic KOH gave j3-sitosterol 4, C29 H SOO (M+
Results and Discussion 4 1 4), mp 1 3 8°C and cerotic acid (hexacosanoic acid)
The ester 1, j3-sitosterol cerotate, mp 93°C ana­ 3, C26Hs202 (M+ 396), mp 88°C (litS mp 88-89°C).
lysed for CSSH O002 , showed in the UV spectrum in Both the products of alkaline hydrolysis were identi­
I
MeOH an absorption at Amax 247 (log E, 3.6 nm) char­ fied by comparison of their I H NMR spectra9 as well
2
acteristic of unsaturated compound . It showed posi­ as by direct comparison (m mp and co-TLC) with au­
tive Liebermann-Burchard colour reaction (pink thentic samples. Therefore, the structure of 1 was elu­
colouration) suggesting its steroid nature. The IR cidated as j3-sitosterol cerotate.
spectrum of 1 showed the presence of ester carbonyl Phenolester 2, C26H43NOs, mp 97°C showed in UV
( 1 735 c m- I ) and olefinic ( 1 635 cm- I ) functions. The spectrum in MeOH absorptions at Amax 258 sh (log E,
400 MHz I H NMR spectrum of 1 in CDCl3 showed 4.04) and 270(4.06) nm and in MeOH+OR at Amax
the presence of two primary methyls (8 0.82,3H,t, 295 nm characteristic of ni trophenols I . The IR spec­
1 = 7.5 Hz, Hr29; 0.84, 3H,t, 1 = 7 .5 Hz, Hr26'), three trum of 2 in KBr pellet showed the presence of in­
secondary methyls (8 0.87, 6H, d, 1 = 6.5 Hz, H 3 -26 tramolecularly bonded hydroxyl (3200 cm' I ) , ester
and 27; 0.92, 3H, d, 1 = 6.5 Hz, H 3 -2 1 ), two tertiary carbonyl ( 1 730 cm' l ) and aromatic nitro ( 1 540 and
methyls (8 0.68, 3 H,s, H 3 - 1 8 ; 1 .02, 3H, s, Hr 1 9), me­ 1 330 cm' l ) functions. The IR bands at 1 625 , 1 580,
thylene and methine protons (8 0.75-2. 1 5 , m), one 1 5 1 8 and 1 460 cm, l also suggested its 1 ,2,4-
methylene adjacent to ester carbonyl (8 2.27, 2H, t, trisubstitutions on the aromatic ring. The 400 MHz I H
1 = 7 .5 Hz), one methine (8 4.62, I H, m, W I /2 1 6.0 Hz, NMR spectrum of 2 in CDCh displayed signals of
H-3) and one olefinic proton (8 5 .37, I H, dd, 1 = 4.5 one methyl (8 0.88, t, 1 = 7.5 Hz), eighteen methyl
 DINDA el aL. : STEROID AND N ITRO PHENOL ESTERS FROM BIGNONIA UNGUIS-CA T! ROOTS 1515

29

�C"'C"'N'"
5
CHJ(CHili4COOH
3

26 l
' CHJ(C�h6COOH
1 R = C�(CH2 )z4CO-
4 R=H 7

18 1
2 R' = CH:3(C�)16CO-
6 R' = H 12 0

�
396 II r=
C H:3(C �h4-C -O� .
+

......
. _.. - 276
CH,(C O-
mJz 424
35 1

Scheme I. Mass fragmentation of 1

1 4. 1 (q, C- 1 8) suggested the presence of stearoyl moi­

enes (& 1 .24, brs, 1 4 X CH2 ; 1 .56, distorted t, J = 7.5
ety. The HMBC experiments (Table I) also con­
Hz, �-CH2 with respect to ester carbonyl ; 2.28, t,
firmed the assignments of proton signals in the alco­
J = 7.5 Hz, -CH 2 adjacent to ester carbonyl; 2.93 , t,
hol moiety of 2. Compound 2 on alkaline hydrolysis
J = 7.5 Hz, benzylic CH2 ; 4.27, t, 1 = 7.5 Hz, -OCHr),
with I N methanolic KOH gave 2-(4-hydroxy-3-
three aromatic protons(& 7 . 1 1 , d, 1 = 8.0 Hz, H-5';
nitrophenyl)ethanol 6 [C gH9N04 (M+ 1 83)] as pale
7 .45, dd, 1 = 8.0 and 2.0 Hz, H-6' and 7.97, d, 1 = 2.0
yellow needles, mp 65°C and stearic acid 7 [C lgH3602
Hz, H-2') and one phenolic hydroxyl proton (& 1 0.49,
(M+ 284)] as granular crystals, mp 69°C (lit8 mp
s, exchangeable with D 2 0, HO-4') suggesting 2-(4-
7 I .5°C). Compound 6 was synthesised from p-cresol
hydroxy-3-nitrophenyl)ethyl ester structure for it. The 2
by methylation 1 with dimethyl sulphate in the pres­
chemical shift values of aromatic protons were as­
ence of anhydrous K2C03 followed by nitration with
signed on the basis of NOE correlation experiments. 2
nitric and sulphuric acids mixture, formylation1 with
The NOE enhancements were observed at & 7.97, 7 .45
formaldehyde gas and demethylation 1 3 of the formy­
and 4.28 when irradiated at & 2.93, while the NOE
lated product (Scheme II). The synthetic product 6
enhancement was only observed at & 2.93 when irra­
was identical (m mp, and co- TLC) with the hy­
diated at & 7 .97 . Similar 'H NMR chemical shift val­ droxynitrophenyl ethanol, obtained from alkaline h � ­
ues were reported for 4-hydroxy-3-nitrophenethyl­ drolysis of 2. Therefore, the structure of 2 was elUCI­
amine 5 isolated from Cereus validus " . The carbon dated as 2-(4-hydroxy-3-nitrophenyl)ethyl stearate.
signals in the 1 00 MHz I 3 C NMR spectrum of 2 at &e The biological activity of the compounds 1 and 2 will
1 73.6(s, C- l ), 34.3(t, C-2), 24.9(t, C-3), 29. 1 (t, C-4), be studied later on. It may be noted that earlier anti­
29.3(t, C-5), 29.4(t, C-6), 29.5 (t, C-7), 29.6(t, C-8), tumour nitrophenanthrene derivative 12 has been re­
29.7(t, C x 7, C-9, 1 5), 3 1 .9(t, C- I 6), 22.7(t, C- 1 7) and ported from A ristoLochia bracteata l 4 .
1516 INDIAN J . CHEM., SEC B , JUNE 2003

I - HMBC NMR data of 2 (alcohol moiety) in CDCI)

Carbon HMBC Carbon HMBC

Table

be be

I' 1 30.4(s) H-2', H-6', H-7' 5' 1 20. I (d) H-6', HO-4'
2' 1 24.7(d) H-6 ', H-7 ' 6' 1 38.3(d) H-2', H-5', H-7'
3' 1 33.4(s) H-2', H-5', H-O-4' 7' 33.9(t) H-2', H-6', H-8'
4' 1 53 .9(s) H-2', H-5', H-6' 8' 63.9(t) H-7'

8 9

11

6
II-Reagents and conditions: (i) (CH)hS04/K2CO) in Me2CO (ii) HNOfH2S04 in ice bath (iii) HCHO gas in THF; CaO (iv)
Separation by CC of IO HI/Red P in AC20, tl..
Scheme

..

Experimental Section column with petrol-C6H6 (4: 1 ) elUate gave a residue

General. All solvents were distilled before use. which on repeated CC afforded 1 in colourless nee­
Silica gels (Merck) were used for column and thin 3
dles (0.03 g, yield, 3.7 x 1 0. %), [Rj, 0.60 in petrol­
layer chromatography . UV, IR, MS and NMR spectra C6H6 (4: 1 ) mixture on silica gel G]. Anal. Found: C,
were recorded on Spectronic 2 1 0, Perkin-Elmer 577, 82.90; H, 1 1 .98. Calc. for C55HIO002: C, 83.26; H,
Jeol JMS-AX 505H and Varian XL 400 spectrome­ 1 2.70%.
ters, respectively. GC was carried out on Pye Unicam Elution of the column with petrol-C6H6 (6:4) gave
1 04 gas chromatograph. Petrol used had the bp 60- a residue which on repeated crystallisation from pet­
80°C. rol-C6� mixture afforded 2 in brown granular crys­
tals (0.025 g, yield, 3 . 1 x 1 0. %), (Rj, 0.45 in C6H6 on
3
Plant material silica gel G). Anal. Found: C, 68.92; H, 9.60; N, 2.74.
The roots of Bignonia unguis-cali were supplied by Calc. for C26H43N05 : C, 69.45 ; H, 9.64; N, 3. 1 2%.
MIs United Chemical and Allied Products, Calcutta in
Alkaline hydrolysis of 1 . Compound 1 (0.01 g)
October 1 999. A voucher specimen has been pre­
served in the herbarium of Shibpur Botanical Garden, was dissolved in 1 0 mL of I N KOH in MeOH and
was refluxed for 2 hr in an atmosphere of N2. The re­
Howrah, West Bengal, India.
action mixture was concentrated and extracted with
Extraction and isolation CHCh ( 1 0 mL x 3). The combined CHCh extract was
Air dried milled roots (0.80 kg) of B. unguis-cali washed with H20, dried over anhydrous Na2S04, con­
were extracted with EtOAc in a Soxhlet-type extrac­ centrated and subjected to Cc. Petrol-C6H6 ( 1 :2) elu­
tor. The extract was concentrated to a residue (3.5 g) ate gave colourless needles of 3, mp 1 38°C (0.003 g).
by distillation under reduced pressure. The residue The aqueous part of the reaction mixture was neutral­
was extracted with CHCh and the CHCh soluble part ised with IN HCI and extracted with Et20 ( 1 0
was concentrated and subjected to Cc. Elution of the mL x 3). The combined ether extract was washed with
 DINDA et al. : STEROID AND NITRO PHENOL ESTERS FROM B1GNON1A UNGUlS-CA Tl ROOTS l S 17

H 20, dried over anhydrous Na2S04 , concentrated and 6.66(2H, d, 1 = 8.0 Hz, H-3,5) , 7 .02(2H, d, 1 = 8.0 Hz,
chromatographed. C6 H 6-CHCl) (6: 1 ) eluate gave 3 as H-2,6).
colourless granular crystals, mp 88°C (0.003 g). Synthesis of 4-methoxy-3-nitromethylbenzene
3: IH NMR (COCI) : 0 0.80(3H, t, 1 = 7.0Hz, CH) , 9. Compound 8 ( 1 .0 g) was dissol ved in 3 mL of
1 .2S(44H, brs, 22 x CH 2 -), 1 .62(2H , quintet, 1 = 7.0 conc. H2S04 in an Erlenmeyer flask and was cooled
Hz, �-CHr), and 2.34(2H, t, 1 = 7.0Hz CH2CO-) in ice-bath. To it 2 mL of conc. HNO) was added
4: I H NMR (COCI) : 0 0.68(3H, s, H3- I 8). dropwise with shaking. After addition of HNO), the
0.8 1 (3H, t, 1 = 7.0Hz, H3-29), 0. 86(6H, d, 1 = 6.S Hz, reaction mixture was kept as such for another I S min
H)-26, 27), 0.92(3H, d, 1 = 6.S Hz, H)-2 1 ), 1 .02(3H, s, and then poured in ice cold H20 (30 mL) and ex­
H)- I 9), 0.7S-2.26(29H, m,-CH2, >CH) , 3.S0( 1 H, m, tracted with CHCh ( 1 0 mL x 3 ) . The CHCh extract
H-3), S.34( l H, brdd, 1 = 4.S and 4.S Hz, H-S ). was washed, dried over anhydrous Na2 S 04 , concen­
Alkaline hydrolysis of 2. Compound 2 (0.0 1 2 g) trated and chromatographed. Elution of column with
was dissolved in I OmL of I N KOH in MeOH and the C 6H6-CHCI) (S: 1 ) gave 9 as brown semi-solid (yield
solution was refluxed for 3 hr in an atmosphere of N2 . 0.70 g , 70%). ' H NMR (COCh) : 0 2.26(3H,s, H3C­
The reaction mixture was concentrated, diluted with I ), 3.82(3H,s, CH30-4), 7 .08( l H,d, 1 = 8 .0 Hz, H-S ),
H20, neutral i sed with I N HCI and extracted with 7.44( 1 H, dd, 1 = 8.0 and 2.0 Hz, H-6), 7 .98( l H , d,
Et20 ( 1 OmL x 3). The combi ned ether extract was 1 = 2.0 H z, H-2).
washed with H 2 0, dried over anhydrous Na2 S04 , con­ Synthesis of 2-(4-methoxy-3-nitrophenyl)ethanol
centrated and chromatographed. Elution of the col­ 10. Compound 9 (O.SO g) was dissolved in THF
umn with C6H6-CHCI3 (6: I ) gave 7 as granular crys­ ( 1 0 mL). To it SO mg of anhydrous CaO was added
tals (0.004 g), mp 69°C. Elution of the column with and HCHO gas was passed by heating paraformalde­
C6H6-CHCI3 (4: 1 ) gave 6 as pale yellow needles hyde for I S min. The cooled reaction mixture was
(O.OOS g), mp 6S°C. fi ltered and washed the residue with a little THF. The
6: UV(MeOH ): 278.8(\ogE, 3.64)nm; IH NMR combined fi ltrate was concentrated and chromatogra­
(COCI3): 8 2.83(2H, t, 1 = 7.S Hz, H2-7), 2.38( 1 H , brs, phed. Elution of the column with C6H6-CHCI) (3: 1 )
exchangeable with 020, HO-8), 3 .92(2H, t, 1 = 7.5 eluate gave 10 as dark brown crystals (yield 0.07g,
Hz, Hr8), 7. 1 O( l H, d, 1 = 8.0 Hz, H-S), 7.46( l H, dd, 14%), mp SS°c. 'H NMR (COCI3): 0 2 .34( 1 H, brs,
1 = 8.0 and 2.0 Hz, H-6), 7 .94( 1 H , ct, 1 = 2.0 Hz, H-2), exchangeable with 020, HO-8), 2.82(2H, t, 1 = 7.0
10.SO( l H, s, exchangeable with 020, HO-4). Hz, benzylic CH2 ), 3.84(3H, s, CH30-4), 3.89(2H, t,
7: IH NMR (COCI3): 0 0.84(3H, t, 1 = 7.S Hz, CH) , 1 = 7.0 Hz,Hr8), 7. 1 4( 1 H, d, 1 = 8.0 Hz, H-S),
1 .2S(28H, brs, 1 4 x CH2), 1 .63(2H , quintet, 1 = 7.S Hz, 7.S0( 1 H,dd, 1 = 8.0 and 2.0 Hz, H-6), 8.02( l H, d,
�-CH2-)' 2.34(2H,t,1 = 7 .S Hz, -CH2CO); ElMS rnIz 1 = 2.0 Hz, H-2). Elution of the column with C6H6-
(%): 284(M+, 20), 2S6(M-28, 4S), 24 1 (M-43, 7), CHCl) (2: 1 ) gave 1 1 (2-methoxy-S-methyl-3-
239(M-4S, 6), 227(M-S7, 8)224(M-60, 7), 7 1 (94), nitrophenyl)methanol as brownish semi-solid (yield
60(S6), S7( 1 00), 43(86). Methyl ester was identical O.2Sg, SO%). ' H NMR (COCI3): 0 2.28 (3H, S , CH3-
with an authentic sample of methyl stearate on GC S), 2.60( l H , brs, exchangeable with 020, -OH),
analysis under the same condition using PEGA col­ 3 .82(3H, s, CH30-2) , 4.82(2H, s, H 2 C- 1 ), 7.S4( 1 H , d,
umn (inj. port, column and FlO temperatures were 1 = 2.0 Hz, H-6), 8 . 1 2( l H, d, 1 = 2.0 Hz, H-4) .
200°C, 1 90°C and 2 1 0°C, respectively ; N 2 at the flow Synthesis of 6. Compound 10 (O.OS g) was dis­
rate SO mL/min and chart speed 320mrnlhr). solved in 3 mL of AC20. To this solution I .S mL of
Synthesis of 4-methoxymethylbenzene 8. p­ H I and SO mg of red P were added and the mixture
Cresol ( 1 .2 g) was dissolved in dry acetone (20 mL). was refl uxed for 2 hr. The cooled reaction mixture
To it S mL of (CH3hS04 and 1 00 mg of anhydrous was fi ltered and the residue washed with a l ittle
K2CO) were added. The resulting mi xture was re­ Ac20 and the combined filtrate was concentrated and
fluxed for 4 hr using CaCl2 guard tube. The cold mix­ diluted with H20 and extracted with CHCh ( 1 0
ture was filtered and the residue was washed with ace­ mL x 3). The CHCl3 extract was washed with H20,
tone. Evaporation of the acetone solution gave an oily dried over anhydrous Na2 S 04 , concentrated and
residue which on column purification afforded 8 chromatographed. Elution of the column with C6H 6-
(yield 1 . 1 g, 92%) as viscous liquid. I H NMR CHCh (2: 1 ) gave 6 as pale yellow crystals (yield
(COCI) : 8 2.2S(3H,s, H3C- 1 ), 3.84(3H,s,CH3 0-4), 0.03g, 60%), mp 6S°C.
1518 INDIAN J . CHEM., SEC B, JUNE 2003

5 Tokes L, Jones G & Djerassi C, J Am Chern Soc, 90, 1968,

5465.
Acknowledgement
6 Tokes L & Djerassi C, J Am Chem Soc, 9 1 , 1969, 5017.
7 Diekmann J & Djerassi C, J Org Chern, 32, 1967, 1 005.
The authors wish to thank Prof. B C Ranu, lACS,

8 Weast R C, Hand Book of Chemistry and Physics, 57th edn,

Kolkata, for I H NMR spectra and RSIC, Lucknow,

(CRC, Ohio), 1976, p c33 1 , 402.

India for microanalysis. One of the authors (U C D) is
9 Pouchert C J & Campbell J R, ALdrich Library of NMR spec­
thankful to the CSIR, New Delhi, India for the award
tra, (Milwaukee, Wisconsin, Aldrich Chern Co.), 1975.
10 Silverstein R M, Bassler G C & Morril l T C, Spectrometric
of a Senior Research Fellowship.

Identification of Organic Compounds, 4th edn, (John Wiley,

1 Rastogi R P & Mehrotra B N, Compendium of Indian Medici­ New York), 1981, p 324.
References

nal Plants, Vol 4, (CSIR, New Delhi), 1995, p 1 04. I I Neme G, Nieto M, D' Arcangclo A T & Gros E G, Phyto­
2 Joshi K C, Singh P & Sharma M C, J Nat Prod, 48, 1985, 1 45. chemistry, 1 6, 1977, 277.
3 Mukherjee K S, Ghosh P K, B hattacharya P & Mukherjee R 1 2 Fieser L F & Fieser M, Reagents for Organic Synthesis, (John
K, J Indian Chem Soc, 64, 1987, 1 29. Wiley, New York), 1967, pp 295, 397.
4 Asahi Research Centre Co, Tokyo, Hand Book of Proton 13 Deulofeu V & Guen·ero T J, Org Syn CoLIn, Vol. 3, 1955,
NMR Spectra and Data, Vol. 5 , (Academic Press, Tokyo), 586.
1986, p 400. 14 Husain F T, PLanta Medica, 1 8, 1969, 30.