CHAPTER - II

THE CONSTITUENTS OF THE LEAVES OF MURRAYA KOENIGII (RUTACEAE).

Introduction 26-33

Results and Discussion 34-42

Experimental 43-47

References 48-51

26

INTRODUCTION

1. CARBAZOLE ALKALOIDS

(a) Carbazole alkaloids containing secondary nitrogen :

Tricyclic : Tricyclic carbazole alkaloids isolated from stem bark of M. koenigii

were mukoeic acid (la) -2, mukonal (lb)^ 1-hydroxy-3-methyl carbazole (Ic)" and

me-2-methoxy carbazole-S-carboxylateCId)". Koenoline (le) isolated from root

bark of M. koenigii exhibited cytotoxic activity against the KB cell-culture test

system^. 2-Methoxy-3-methyl carbazole (If)^ from stem bark and murrayanine

(ig) ' from leaves and stem bark of M. koenigii yNere also isolated. Other tricy­

clic carbazole alkaloids isolated from M. koenigiiwere mukonidine (Ih)^, mukoline

(li)^°, mukolidine (Ij)'", mukonine (lk)^\ 2-hydroxy-3-methyl carbazole (ll)^^ 3-

methyl carbazole (Im)", glycozoline^^ mukoenine-A, -B and -D^^ The tricyclic

carbazole alkaloids having branched side chain, isolated from the stem bark

and seeds of M. koenigii, were grinimbilol (li)^^ mahanimbinol = mahanimbilol

(liia)«' ^ and murrayanol (lllb)^^

Tetracyclic : Murrayacine (iVa)"'^^ and koenidine = koenigicine = koenimbidine

(IVb) -i9-2i from leaves and roots, grinimbine (IVc) ^^^ from stem bark, leaves

and roots, koenimbin = koenimbine (IVd^^'^from fruits, mukonicine {IVe) '*from

leaves along with koenine (IVf) ^^^ and koenigine (IVg) ' ^ were isolated from

M. koenigii having no side chain. However tetracyclic carbazole alkaloids

containing branched side chain included mahanimbine {Way'*^-^^-^^^^ irom fruits

and leaves, mahanimbicine = isomahanimbin (Vb) - - from roots and leaves,

isomahanine (Vc)'^ from seeds, mahanine (Vd)^^2^3° from leaves and

27

I

a

b

c

d

e

f

9

h

i

i

k

1

m

R2 = R''

R^ = R^

R2 = R4

R^ = R^

R2 = R''

R^ = R''

R2 = R'

R^ = R^

R2=R3

R2 = R3

R2 = R''

R^ = R^

Ri = R2

H, R = OCH3, R3 = CO^H

H, R2 = 0 H , R3 = C H 0

H, R = OH, R3 = CH3

H, R2 = OCH3, R3 = CO2CH3

H, R = OCH3, R3 = CH^OH

H, R2 = OCH3, R ' = CH3

H, R = OCH3, R^ = CHO

H, R2 = OH, R3 = CO2CH3

: H, R = OCH3, R" = CH^OH

: H , R = OCH3, R^ = CHO

: H, R'= OCH3, R3 = CO2CH3

: H, R2 = OH, R3 = CH3

: R'' = H, R3 = CH,

II

28

a R2 = R3 = H, Ri = C K

b Ri = H, R2 = CH„ R ' = OH

^ ^ ^ ^ % , ^

IV

a Ri = R3 = R" = H, R2 = CHO

b R = H, R = CH,, R2 = R3 = OCK

c R2 = R3 = R4 = H, R = CH,

d R3 = R" = H, R =CH,, R2 = OCH,

e R3 = H, Ri = CH„ R2 = R* = OCH,

f R3 = R4 = H, R = CH,, R2 = OH

R'' = H, R = CH„ R = OCH,, R = OH

29

a Ri = R2 = R3 = H

b Ri = R3 = H, R2 = CH3

C Ri = H, R2 = CH3, R3 = OH

d R2 = H, R = CH3, R3 = OH

(CHJXH(OH)

C(CH3):CH,

VI

Vil

30

VIII

a R = C(CH3)pH

b R = C(CHJCH,

a

b

R = H

R = CH,

31

mahanimboline (Vl)^^ were also isolated from M. koenigii.

Pentacyclic : The only pentacyolic carbazole alkaloid without any side chain

isolated from stem bark of M. koenigii was murrayazolidine (Vn)^^ while

murrazolinine (VlllaP and cyclomahanimbin = currayanine (Vlllb)"*' isolated

from leaves, possess branched side chain.

Hexacyclic : Bicyclomahanimbicine (IXa)^from leaves and bicyclomahanimbin

(rXb) '2^ from both leaves and stem bark of M. koenigii v^ere isolated.

Carbazole alkaloids murrayanine, grinimbine and mahanimbine

containing secondary nitrogen isolated from stem bark of M. koenigiia\so showed

antifungal activity against some human pathogenic fungii^^.

(b) Carbazole alkaloids containing tertiary nitrogen :

Isomurrayazoline (Xa)^^, currayangine = mahanimbidine = murrazoline

(Xb)^2S2^** from leaves and stem bark while murrazolinol (Xc)^^ from stem bark

of M. koenigiiyNere isolated. The crystallographic parameters for murrayazoline^^

are a 11.19, b 11.05, c 15.65 e, p 115.27°, d. (exptl.) = 1.24, z = 4, d. (calcd.) =

1.258, space group P21/c, R = 0.074.

(c) Binary carbazole alkaloids and quinones :

Binary carbazole alkaloids and quinones isolated from M. koenigii

included murrastifojine - F, bis-2-hydroxy-3-methyl carbazole, bismahanine,

biskoeniquinone-A and bis murrayaquinone-A^".

2. ESSENTIALOILS

a- pinene^^"", l-caryophylene^^, sabinene^^ P-caryophyllene'*\ p-

elemene'*\ p-phellandrene^V terpenen-4-ol' '° from leaves while 8-

HsC

a R = H, R' = H

b R = CH3, R' = H

c R = CH„ R' = OH

H3ca

GluO

XI Glu = glucopyranosyl

/SD

XII

CH„OH r^/T^O' 2 ' OCMGg

. / ^ O

H H

XIII

33

geranyloxypsoralene''2, imperatorin''^, heraclenin''^, isosaxalin''^ and

heraclenoFwere isolated from the seeds of M. koenigii.

3. MISCELLANEOUS

Glycoside scopoline (Xl)^ glucose^ from leaves and 3-(1,2-dimethyl

allyl) xanthyletin (Xll)"^ from stem bark of M. koenigii were isolated. Marmesin-

1"-0-p-D-galactopyranoside (XIII) and umbelliferone isolated from stem bark of

M. koenigii showed antiinflammatory and antimicrobial activities'".

34

RESULTS AND DISCUSSION

The dried leaves of Murraya koenigii were extracted with methanol and

methanol extract was concentrated and treated with petrol, benzene, ethyl ac­

etate and acetone. The petrol fraction on column chromatography using silica

gel as adsorbent and eluting with petrol-benzene (1:1), yielded a crystalline

product MK-1.

MK-1, crystallized from benzene, m.p. 205°C, gave brown fluorescence

in UV light after developing on thin layer chromatography. In the infra red spec­

trum bands appeared at 3400 (N-H stretching), 1640, 1610 and 1580 cm-

(unsaturation and aromatic system). It analysed for C^gH jgNOg (M* 293). In the

mass spectrum the molecular ion appeared at m/z 293 and base peak appeared

at m/z 278 (M-CH3). Other prominent peaks appeared at m/z 235 (M-58) and

m/z 139. The odd mass number for molecular ion is indicative of the presence

of nitrogen.

In the 'H NMR spectrum (600 MHz, CDCI3) given in Table 1 and Fig.1, a

singlet integrating for six protons at 8 1.485 can be assigned to gem dimethyl

protons. The singlet at 5 2.340 is attributable to benzylic methyl group while a

singlet at 5 3.903 can be assigned to methoxy protons. A pair of doublet

appearing at 5 5.640 (J=10 Hz) and 6.536 (J=10 Hz) were assigned to 3'-H and

4'-H, respectively, of chromene ring. The spectrum shows signals for aromatic

protons at 5 7.627 (s, 4-H), 6.940 (dd, J=8.4. 2.2 Hz, 7-H), 7.214 (d. J=8.4 Hz,

8-H) and 7.421 (d, J=2.2 Hz, 5-H). The broad singlet at 6 7.686 was assigned to

N-H proton. The assignments were supported by DEPT experiments (Fig. 2)

and HHCOSY correlations (Fig. 3).

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35

The "C NMR spectrum (150.8 MHz, CDCg given in Table I and Fig.

4,shows signals for all the 19 carbons. The signal at 516.022 was assigned to

methyl carbon C-3 of carbazole nucleus which was supported by DEPT experi­

ments. The geminal methyl group carbons at C-2' absorb at 6 27.596 while

methoxy carbon attached to C-6 gives signal at 556.038 and C-2' absorb at 5

75.849 which was confirmed by DEPT experiments. The ^ C NMR signals for

sp^ carbons in the carbazole skeleton are as follows 5102.680 (C-5 ), 104.505

(C-1), 111.048 (C-8), 113.021 (C-7), 116.868 (C-4b), 117.197 (C-4'), 118.299

(C-3). 121.028 (C-4), 124.448 (C-4a). 129.199 (C-3'), 134.410 (C-8a), 135.759

(C-9a), 149.832 (C-2), 153.975 (C-6). The assignments were further confirmed

by DEPT experiments, HMBC (Fig. 5), COLOC (Fig.6), and CHCOSY (Fig.7)

correlations. On the basis of above data MK-1 was confirmed as koenimbine

(IVd) m.p. 205° C (lit. 194-95° C) ^^^ which was characterized earlier on the

basis of ^H NMR, IR, UV and MS only. The ^^c NMR and 2D NMR is discussed

for the first time.

IVd

Further elution of the column with petrol - benzene and crystallization

from benzene gave colourless crystals of MK-2.

MK-2, m.p. 139-40°C was UV inactive and it was detected by charring of

TLC plate (SiOg) with perchloric acid. In the IR spectrum a broad band at 3400

36

Table I. 'H NMR and '^C NMR data of MK-1 (IVd)

Assignment

C/H

Chemical Shift (S.ppm)

13 C NMR (150.8 MHz, CDCL) 'H NMR (600MHz, CDCL)

1

2

3

4

4a

4b

5

6

7

8

8a

9

9a

3-Me

6-OMe

1'

2'

3'

4"

2'-Me.

104.505

149.832

118.299

121.028

124.448

116.868

102.680

153.975

113.021

111.048

134.410

-

135.759

16.022

56.038

-

75.849

129.199

117.197

27.596

7.627 (s)

7.421 (d, J=2.2 Hz)

6.940(cld,J=8.4,2.2 Hz)

7.214(d, J=8.4Hz)

7.686 (br,s)

2.340 (s)

3.903 (s)

5.640 (d,J=10Hz)

6.536 (d,J=10 Hz)

1.485 (s)

37

cm^ indicated the presence of -OH group and the other bands were compa­

rable with the IR spectrum of authentic sample of p-sitosterol"^. The occur­

rence of p-sitosterol from M. koenigii has not been reported so far.

Elution of the column with benzene followed by preparative thin layer

chromatography yielded MK-3 and MK-4.

MK-3 was crystallized as colourless granular compound from alcohol -

benzene (7:3) melting at 94.5°C. It was UV inactive and its homogeniety was

checked on TLC plate (SiOg) with iodine as well as perchloric acid. In the mass

spectrum, molecular ion appeared at m/z 476 with successive loss of 14 show­

ing peaks at m/z 462, 448, 434, 406 etc. and base peak appearing at 57, indi­

cating the presence of long chain hydrocarbon.

The ^H NMR spectrum (600 MHz, CDCy is given in Fig. 8. It shows

signals at 8 0.866 (t, J=6.6 Hz), 1.243 (s), 1.542 (d, J=5.88) and 3.618 integrat­

ing in the ratio 3 : 66 : 3 : 2 respectively. The consideration of ^H NMR integra­

tion of 74 protons and the signal at 5 3.618 for two protons can lead to a

molecular ion C^^H^JD (M^522), but the molecular ion peak at m/z 522 is not

indicated in MS. In MS, a fairly unstable peak appeared at m/z 476 (3.5) as

molecular ion peak.

The ^^C NMR spectrum (150.8 MHz, CDCI3), given in Fig. 9, shows sig­

nals at 6 14.065 (-CH3), 22.680 (-CH^-), 25.771 (-CH^-), 29.355 (-CH^-), 29.454

(-CH^-), 29.618 (-CH^-), 29.700 (-CH-)^, 31.936 (-CHg-), 32.873 (-CH^-) and

63.108 (-CH^O-). The appearance of ^ C NMR signal at 5 63.108 and proton

signal at 5 3.618 indicates the presence of CHg group attached to oxygen,

probably an alcohol. The assignment of CH^and CH3 carbons has been

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MK-4 crystallized from chloroform, m.p. 235-37°C, C gH gNO (M^ 331),

gave brown fluorescence in UV light after developing on TLC plate (silica gel.)

The IR spectrum shows bands at 1640 and 1600 c m \ characteristic of

unsaturation.The ^H NMR spectrum (Fig.11) in CDCI3 at 100 MHz was compa­

rable with that of mahanimbidine^ and shows singlets for methyl groups at 6

1.32, 1.48, 1.92 and 2.34 attributable to 8'-Me, 3'-Me, 9'-Me and 3-Me, respec­

tively (Table II). The muJtiplets appearing at 8 0.20 and 1.40 were assigned to

5'-ax-H and 5'-eq-H, respectively. The benzylic proton at 1'- position appears

as a broad doublet at 5 3.35. The aromatic proton at C-4 appears as a singlet at

5 7.35. Other aromatic protons absorb between 6 6.30 - 7.90 as double dou­

blets or multiplets integrating for four protons at 5, 6,7and 8 carbons. Although

chemical and spectral data of MK-4 (m.p. 234-37°C) are comparable with

mahanimbidine (Xb)^ (scurrayangine = murrazoline)" 7 34 p p 266°C, yet

Table II. 'H NMR data of compound MK-4 (100 MHz, CDCi,)

Assignment Chemical shift ( 5, ppm)

5'-ax-H 0.20 (m,1H)

8'-Me 1.32 (s,3H)

5'-eq-H 1.40 (m,3H)

3'-Me 1.48 (s,3H)

9'-Me 2.34 (s, 3H)

3-Me 2.34 (s, 3H)

1'-H 3.35(brd, 1H)

Ar - CH3 7.35 (s, 3H)

Ar-H 6.30-7.90 (m, 5H)

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39

its structural assignment is not conclusive due to large difference in melting

point. Further studies are required to settle the stereochemistry as well as struc­

ture.

The benzene fraction on column chromatography (silica gel), eluting the

column with petrol - benzene (1:1) and benzene - ethyl acetate (8:2) gave

Mk-5andMK-6.

MK-5 was obtained as light yellowish green crystals from benzene - al­

cohol. It melts at 225°Cand shows brown fluorescence in UV light after devel­

opment on TLC plate (silica gel). The IR spectrum shows bands at 3400 (N-H),

1640, 1620 and 1600 cm" (unsaturation and aromatic system). In the mass

spectrum molecular ion appeared at m/z 323 and base peak at m/z 308.The ^H

NMR spectrum (Table III and Fig. 12) was comparable with that of koenldlne

(IVb)^^ (=Koenigicine = Koenimbldine)^^ 21 n p 224-25°C. The structure was

confirmed by ^^c NMR (Table III and Fig. 13), DEPT experiments, HHCOSY

(Fig. 14), CHCOSY (Fig.15), COLOC and HMBC (Fig. 16) studies which was

not done earlier.

CH3O

CH3O

IV b

MK-6 was obtained as light green granules from pyridine - benzene -

methanol (1:11:10). It melts at 288 - GO C (lit. 2Q0°CY^ and was detected by

iodine and perchloric acid on TLC plate (silica gel.). In the infra red spectrum it

40

Table III. 'H NMR and ' 'C NMR data of MK-5 (iVb)

Assignment

C/H

Chemical shift ( 5 , ppm)

13 C NMR (150.8 MHz, CDCL) ^H NMR (600 MHz, CDCL)

1

2

3

4

4a

4b

5

6

7

8

8a

9

9a

3-Me

6-OMe

7-oMe

2'

3'

4'

2'-Me,

104.620

148.681

118.282

120.124

124.448

116.868

102.286

148.188

144.390

94.723

134.410

-

135.759

16.022

56.301

56.581

75.668

. 129.429

117.263

27.547

7.532 (s)

7.367 (s)

6.874 (s)

7.726 (s)

2.311 (s)

3.902 (s)

3.957 (s)

5.657 (d,J=10Hz)

6.570 (cl,J=10Hz)

1.463 (s)

41

shows strong absorption at 3350 cm"" indicating the presence of -OH group. Its

melting point and IR spectrum indicated it to be sterol glycoside. The structure

was assigned as sitosterol - 3 - 0 - p - D - glucoside (XlVa) from IR and H

NMR (100 MHz, CDCI3) given in Fig. 17 and Table IV of its acetylated derivative

(XlVb). The occurrence of sitosterol - 3 - O - P - D - glycoside from M. Koenigii

has not been reported so far.

OR'

OR'

XIV

aR' = H

b R' = Ac

42

Table IV. H NMR data of MK-6 tetraacetate (100 MHz, CDCI3)

Assignment Chemical shift ( S , ppm)

18-Me 0.68 (s,3H)

28 - Me 0.76 (d, J=6.8 Hz, 3H)

0.84 (d, J=6.5 Hz. 6H)

0.90 (d, J=6.5 Hz, 3H)

1.00 (s,3H)

1.88-2.32(8, 12H)

3.22 - 3.80 (m, 8H)

5.40 (m, 1H)

1.12 - 1.60 (cyclic & side (chain)

26, 27

21-Me

19-Me

-Me

-OCO Me

-CH,0-

=CH

-CH2, • •CH

43

EXPERIMENTAL

The dried leaves (1.5 kg) of M. koenigii were extracted with methanol.

Repeated extractions were made until the solution become colourless. The

methanol extracts were mixed and dried under reduced pressure. A dark green

gummy mass (100g ) was obtained which was treated with petrol, benzene,

ethyl acetate and acetone several times till solvent become colourless in each

case.

The petrol fraction (40g) was adsorbed on silica gel (30g) and trans­

ferred to column of silica gel (150g) set with petroleum ether (60-80°C). The

elution of column with petrol - benzene (1:1) gave crude MK-1 which on disso­

lution in benzene - chloroform, crystallization and recrystallization gave pure

MK-1.

MK-1: Crystalline compound (400 mg), crystallized in benzene which melts at

205°C and dissolves readily in chloroform. Rf=0.2958 (silica gel, petrol- ben­

zene, 4:6) , in UV light gave brown fluorescent, adsorb iodine, developed on

charring with perchloric acid and with alcoholic FeCl3 gave yellowish green

colouration.

UV, IR, MS and 'H NMR data of MK-1.

UVX^*°'^nm (log e) : 360 (4.20), 340 (4.24), 300 (4.62), 240 (4.69), 230 (4.68). m3x

IRv*^^'cm-^ : 3400, 2950, 2850, 1640, 1610, 1580, 1490, 1450, 1420, 1400, max

1380, 1360, 1330, 1290, 1260, 1240, 1200, 1170, 1130, 1120, 1100,

1050, 1020, 890, 870, 840, 800, 770, 760, 720, 680.

MS m/z (rel. int.) : 293 [M]* (40), 278 [M-15] (100) aromatlzation of chromene

ring by loss of Me group, 262 (10), 235 (22), 191 (10), 139(16), 117(10),

77(10), 53(18).

44

^H NMR (600 MHz ) : 5 1.485 (s, 6H, gem dimethyl protons), 2.340, (s, 3H, 3-

Me), 3.903 (s, 3H, Ar-OCHg), 5.640 (d, J=10 Hz, 3'-H), 6.536 (d, J=10Hz,

4'-H), 6.940 (dd, J=8.4, 2.2 Hz, 7-H), 7.214 (d, J=8.4 Hz, 8-H), 7.421 (d,

J=2.2 Hz, 5-H), 7.627 (s, IH, 4-H), 7.686 (br s, IH, N-H).

^ C NMR data is given in discussion (Table I).

From UV, MS, IR, 'H NMR and ^ C NMR spectral data the compound was

found to be koenimbine = koenimbin.

Further elution of column with petrol - benzene (1:1) and crystallization

in ethanol - chloroform (8:2) crude MK-2 was obtained.

MK-2 : Crude MK-2 on crystallization in benzene gave colourless crystals (180

mg) of Mk-2 (pure), readily dissolved in chloroform and having melting point

139-41 °C. Rf=0.7384 (silica gel, benzene - acetone, 9:1) . UV inactive but ap­

peared on absorption with iodine and charring with perchloric acid.

IR, MS and 'H NMR data of MK-2.

IRv l^ax^m ••• : 3400, 3250, 2900, 2850, 1640, 1370, 1050, 950, 880.

MS m/z (rel. int. ) : 414 [M]* (100), 399 (25), 396 (30), 381 (18), 273 (50). 271

(44), 255 (60), 231 (30), 213 (50).

^H NMR (100 MHz) : 5 0.68 (s, 3H, 18-Me), 0.75 (d, J=6.8 Hz, 3H, 28 -Me), 0.84

(d, J=6.5 Hz, 6H, 26, 27-Me), 0.92 (d, J=6.5 Hz, 3H, 21 -Me), 1.01 (s, 3H,

19-Me), 1.07 - 2.34 (-CH^ and -CH protons of cyclic and side chain), 3.52

(m, IH , 3-ax-H), 5.15 (m, IH, -OH), 5.39 (m, IH, oleflnic proton).

From IR, MS, ^H NMR spectral data and on direct comparison with authentic

sample the compound was found to be p- sitosterol.

Elution of petrol fraction column with benzene, applying preparative thin

45

layer chromatography (PTLC) and on crystallization MK-3 was isolated.

MK-3 : Colourless granular compound (120 mg), soluble in chloroform, crystal­

lized in alcohol - benzene (7:3) having melting point 94-5°C. Rf= 0.6484 (silica

gel, benzene, PTLC), UV inactive, absorb iodine and developed on charring

with perchloric acid.

Spectral data of MK-3.

IR V *^ '' cm- : 3300, 2900, 2820, 2300, 1460, 1420, 1370, 1330, 1060, max

1010,760,720. •

MS m/z (rel. int.) : 476 [M]* (3.5), 462 (1.2), 448 (7.5), 434 (1.4), 420 (5), 406

(1), 181 (2), 167 (3), 153 (4), 139 (10), 125 (19), 111 (36), 97 (66), 83

(69), 69(56), 57(100).

^H NMR and ^ C NMR is given in discussion.

From IR, MS, ^H NMR and ^ C NMR the compound was found to be a long

chain alcohol.

Further elution of column with benzene, crystallization and recrystalliza-

tion in benzene MK-4 was isolated.

MK-4 : Colourless, crystalline compound (13 mg) which dissolves readily in

chloroform and melts at 235 - 3 7 ^ . Rf = 0.8238 (silica gel, petrol - benzene,

4:6), in UV light gave brown fluorescent, absorb iodine and developed on char­

ring with perchloric acid.

Spectral data of MK-4 :

UV>.^^°^(log e) 241 (4.62), 257 (4.37), 307 (4.20), 355 (3.51) iTl3X

IRv'^^'cm-^ : 2950, 2900, 1640, 1600, 1490, 1450, 1370, 1350, 1330, 1300, max

46

1250. 1220, 1200, 1180, 1150, 1120,1080, 1060, 990, 850, 740.

MS 70 eV, m/z : 331 [Mr, 316 [base peak], 248.

H NMR (100 MHz) : 5 0.20 (m, 1H, 5'-ax-H), 1.32 (s, 3H, 8'-Me), 1.40 (m,

1 H,5'-eq-H), 1.48 (s, 3H,3'-Me), 1.92 (s, 3H, 9'-Me), 2.34 (s, 3H, 3-Me),

3.35 (br d, 1H, benzylic proton at 1"), 7.35 (s, 3H, Ph - CH3), 6.30 - 7-90

(m, 5H, Ar-H).

From UV, IR, MS and ^H NMR spectral data the compound was found to be

mahanimbidine = currayangine = murrazotine.

The benzene fraction (18g) adsorbed on silica gel (15g) and transferred

to the silica gel column set with petroleum ether (60-80''C). Elution of column

with petrol - benzene (1:1) and crystallization in alcohol - benzene (8:2) MK-5

was isolated.

MK-5 : Light yellowish green crystals (600 mg) melting point 225°C and soluble

in chloroform. Rf=0.3584 (SiOj, benzene), in UV light gave brown fluorescent,

adsorb iodine, developed on charring with perchloric acid and with alchoholic

FeClg gave green colouration.

Spectral data of MK-5 .

^j^^EtOH ^i^g gj . 235 (4 5g)_ 240 (4.57) , 288 (4.36), 297 (4.53). 336 (3.97).

IRv^^'cm-^ : 3400, 2900, 2800, 1640, 1620, 1600, 1585, 1490, 1470, 1455, max . I . . . , . I .

1440,1420, 1400,1370,1355, 1340,1290, 1270,1240, 1200, 1185, 1170,

1150, 1120, 1110, 1050, 1030, 990, 970. 940, 920, 860, 820, 760, 740,

720, 670.

MS m/s (rel. int.) : 323 [M]^ (60), 308 [M-15] (100) aromatization of chromene

ring by loss of methyl group, 293 [M-30] (20) further loss of methyl group.

47

264 (10), 250 (16), 154 (14). 133 (10), 71 (16).

^H NMR (600 MHz) : 5 1.463 (s, 6H, 2'-Me2), 2.311 (s, 3H, 3-Me), 3.902 (s, 3H,

6-OMe), 3.957 (s, 3H, 7-OMe), 5.657(d, J=10Hz, 1H, 3'-H), 6.570 (d, J=10Hz,

1H, 4'-H), 6.874 (s, 1H, 8-H), 7.367 (s, 1H, 5-H), 7.532 (s, 1H, 4-H).

"C NMR is given in discussion (Table III). From UV, IR, MS, ^H NMR and ^ C

NMR the compound was found to be koenimbidine s koenigicine = koenidine.

Further elution of column with benzene - ethyl acetate (8:2) and on crys­

tallization in pyridine - benzene - methanol (1:11:10) MK-6 was isolated.

MK-6 : Light green granules (105g), soluble in pyridine and boiling methanol,

melt at 290°C. Rt= 0.6421 ( SiO^, ethyl acetate - methanol, 8:2 ), UV inactive,

adsorb iodine and developed on charring with perchloric acid.

IR data of MK-6 :

IR\) 'cm- : 3350, 2950, 2850, 1460, 1430, 1380, 1370, 1160, 1110, 1070. max

1010, 950, 920, 880.

Acetylation of MK-6: A tetra acetate (m.p. 158°C) was prepared by treatment

of MK-6 with A c p and pyridine. Rf=0.900 (SiOg", benzene - acetone, 7:1). de­

veloped by adsorption of iodine and charring with perchloric acid.

IR\)'^^'cm-^: 2900. 2850, 1750, 1740, 1460, 1430, 1370, 1210, 1150,1030,900 max

H NMR (100 MHz) : 6 0.68 (s, 3H, 18-Me), 0.76 (d. J=6.8 Hz. 3H, 28-Me), 0.84

(d, J=6.5 Hz, 6H, 26. 27-Me). 0.90 (d, J=6.5 Hz. 3H. 21-Me). 1.00 (s, 3H. 19-

Me). 1.88-2.32 (s, 12H, - OCO CH3), 3.22 - 3.80 (m, 8H.-CHp-). 5.40 (m, 1H.

=CH), 1.12 -1.60 (-CHg, - CH protons of cyclic and side chain).

From IR of MK-6, IR and ^H NMR of its tetraacetate derivative the compound

was found to be sitosterol - 3 - 0 - p - D - glucoslde.

48

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