steroids 7 3 ( 2 0 0 8 ) 579–584
581
26). 13C NMR (400 MHz, CDCl3: ␦ 142, 141 (pyrazine carbons),
2.2.7. (25R)-4-en-spirosta-3,6-dione (9)
Freshly prepared Jones reagent was added dropwise to a solu-
tion of 2 (2.0 g, 4.82 mmol) in acetone (150 mL) at 10 ◦C. The
reaction mixture was stirred below 20 ◦C for 30 min. The reac-
tion mixture was extracted with brine solution (2 mL × 20 mL),
dried over NaSO4, concentrated. The residue was purified by
column chromatography 20% (ethyl acetate/hexane) to give a
white solid of pure 9. Yield: 1.95 g (95%), lit. mp 190–192 ◦C [21].
IR (KBr, cm−1): 2955, 1687, 1610 and 1456. 1H NMR (400 MHz,
CDCl3): ı 6.3 (s, 1H, C4), 4.36 (dt, 1H, C16-␣H), 3.45 (m, 2H, C26),
3.37 (t, 2H, C26). 13C NMR (400 MHz, CDCl3): ı 202 (C3), 199 (C6),
163 (C5), 127 (C4), 109 (C22), 80.6 (C16), 67 (C26), 62.7 (C17), 56
(C14).
203 (C6), 109 (C22), 81 (C16), 67 (C26), 63 (C17). Anal. Calcd. for
C55H80N2O6: C, 76.35; H, 9.32; N, 3.24. Found: C, 75.68; H, 8.97;
3.
Results and discussion
cholestane skeletons connected at the C-2 and -3 positions by
two nitrogen atoms to form a pyrazine ring. They have been
synthesized by the dimerization of two ␣-amino ketosteroids
[14,15]. Initially, the synthesis of dimer 5, which has a double
bond at C5-6, was carried out, as shown in Scheme 1. Oxidation
of diosgenin (2) with PCC to give the known ketone 3 [17]. The
next step was to introduce an amino group or its equivalent (N3
or NO2) at C2 of this ketone. This aim was achieved by bromi-
nation of the carbonyl group followed by substitution of the
bromide by an azido group. Therefore, selective bromination
of ketone 3 by phenyl trimethylammonium perbromide (PTAB)
gave the ␣-bromo ketone 4 as the thermodynamic product.
The 1H NMR spectrum for product 4 shows doublet of doublet
peak at 4.5 ppm for  proton at C2 and doublet of triplet peak
at 4.36 ppm for one ␣ proton at C16. The 13C NMR spectrum
shows a new peak at 204 ppm for carbonyl carbon, 121 ppm
for C6 and a peak at 54 ppm for C2.
2.2.8. (25R)-4-en-2˛-bromo-spirosta-3,6-dione (10)
Procedure as described for 4 afforded a white solid of 10. Yield:
0.16 g (67%), mp 187–190 ◦C. IR (KBr, cm−1): 2946, 1680, 1611,
983. 1H NMR (400 MHz, CDCl3): ı 6.3 (s, 1H, C4), 4.47 (dt, 1H,
C16-␣H, J = 6.7, 6.3), 4.44 (d, 1H, C2-H, J = 3.6), 3.46 (m, 2H, C26),
3.37 (t, 2H, C26, J = 10.8 Hz). 13C NMR (400 MHz, CDCl3): ı 198
(C3), 195 (C6), 159 (C5), 129 (C4), 109.5 (C22), 80.6 (C16), 67 (C26),
62.7 (C17), 57 (C14). Anal. Calcd. for C27H37BrO4: C, 64.15; H,
7.38. Found: C, 63.79; H, 7.10.
2.2.9. Di(25R-6 oxo-4-en-spirostano[2,3-b:2ꢀ,3ꢀ-e])
pyrazine (11)
The dimerization of the C-5 unsaturated-2␣-bromoketone
4 involved two steps to achieve the formation of the pyrazine
dimer 5. The first step is the substitution of bromo atom with
an azido group using sodium azide in the presence of cat-
alytic amount of sodium iodide. This keto azide is not stable at
room temperature and it easily converts into enamino ketone
(hygroscopic) which failed to dimerize under different con-
ditions. Therefore, the azido intermediate was not isolated
but it was reduced to an ␣-amino ketone by triphenylphos-
phine in dry THF followed by addition of water to hydrolyzed
the aza-wittig intermediate. The TLC of the reduction pro-
cess afforded a mixture of products. Thus, the intermediate
␣-amino ketone was not isolated. It was subjected to dimeriza-
tion conditions by stirring in ethanol and toluene containing
a catalytic amount of toluenesulfonic acid and open to the
atmosphere to facilitate aromatization to yield the C-5 unsat-
urated pyrazine dimer 5 in good overall yield. The 1H NMR
spectrum shows a triplet peak at 5.4 ppm for C6 proton and
a doublet of triplet peak at 4.4 ppm for one ␣ proton at C16.
The 13C NMR spectrum shows two peaks at 143 and 144 ppm
characteristics for the pyrazine carbons.
Since cephalostatin 1 has a saturated ring attached on
either side of the pyrazine ring, the next target structure was
the saturated symmetrical bis-steroidal pyrazine dimer 8. This
dimer lacks the double bond at C5. Therefore, hydrogena-
tion of ketone 3 using palladium on carbon afforded product
6 in excellent yield. In the 1H NMR spectrum the peak at
5.3 ppm which was characteristics for the C6 hydrogen dis-
appeared which was in good agreement with literature [18].
Selective bromination of ketone 6 by phenyl trimethylammo-
nium perbromide (PTAB) furnished ␣-bromo ketone 7 as the
thermodynamic product. The 1H NMR spectrum of 7 shows
a doublet of doublet peaks at 4.73 ppm (J = 6, 7.2 Hz) for the
 proton at C2, and the 13C NMR spectrum shows the car-
Procedure as described for 8 afforded pure dimeric pyrazine
11. Yield: 0.11 g (68%), mp 300 ◦C (d). IR (KBr, cm−1): 2922, 1684,
1607 and 1416. 1H NMR (400 MHz, CDCl3): ı 6.0 (1H, s, H4), 4.4
(1H, dt, 16␣), 3.4 (2H, m, H-26). 13C NMR (400 MHz, CDCl3: ı 146,
145 (pyrazine carbons), 202 (C6), 161 (C5), 129 (C4), 109 (C22), 81
(C16), 67 (C26), 62 (C17). Anal. Calcd. for C55H76N2O6: C, 76.71;
H, 8.89; N, 3.25. Found: C, 76.25; H, 8.77; N, 3.00.
2.2.10. (25R)-5˛-spirosta-3,6-dione (12)
A solution of 9 (1.0 g, 2.35 mmol) in ethyl acetate (30 mL) was
stirred with 10 wt% palladium on carbon (10 mg) under hydro-
gen at 40 ◦C for 2 days. The reaction mixture was filtered
over a bed of celite, the solvent was evaporated in vacuum.
Recrystallization in methanol afforded 12. Yield: 0.8 g (80%),
mp 230–231 ◦C, lit. mp 230–232 ◦C [21]. IR (KBr, cm−1): 2960,
1708, and 1706. 1H NMR (400 MHz, CDCl3): ı 4.67 (1H, dd, H2),
4.36 (1H, dt, 16␣), 3.45 (1H, m, H-26), 3.37 (1H, t, H26␣). 13C
NMR (400 MHz, CDCl3): ı 204 (C3), 200 (C6), 109 (C22), 81 (C16),
67 (C26), 62 (C17).
2.2.11. (25R)-2˛-bromo-5˛-spirosta-3,6-dione (13)
Procedure as described for 4 gave a white solid of bromodione
13. Yield: 0.35g (74%), mp 184–187 ◦C. IR (KBr, cm−1): 2965, 1686
and 1705. 1H NMR (400 MHz, CDCl3): ı 4.72 (1H, dd, H2), 4.45
(1H, dt, 16␣), 3.46 (1H, m, H-26), 3.37 (1H, t, H26␣). 13C NMR
(400 MHz, CDCl3: ı 205 (C3), 201 (C6), 109 (C22), 81 (C16), 67 (C26),
63 (C17). Anal. Calcd. for C28H43BrO4: C, 64.24; H, 8.28. Found
C, 63.88; H, 8.15.
2.2.12. Di(25R-6 oxo-5˛-spirostano[2,3-b:2ꢀ,3ꢀ-e]) pyrazine
(14)
Procedure as described for 5 afforde pure dimeric pyrazine 14.
Yield: 0.2 g (78%), 280 ◦C(d). IR (KBr, cm−1): 2950, 1705 and 1407.
1H NMR (400 MHz, CDCl3): ı 4.4 (1H, dt, 16␣), 3.4 (2H, m, H-