Preparation and characterization of novel 4-bromo-3,4-dimethyl-1-phenyl-2- phospholene 1-oxide and the analogous phosphorus heterocycles or phospha sugars

Article abstract of DOI:10.1016/j.bmcl.2010.01.061

4-Bromo-3,4-dimethyl-1-phenyl-2-phospholene 1-oxide (3c) was first synthesized from 3,4-dimethyl-1-phenyl-2-phospholene 1-oxide (2c) by a bromo-radical substitution reaction occurred at C-4 position by N-bromosuccinimide and 2,2′-azobisisobutyronitrile. The novel phospha sugar analogue 3c exerted high anti-proliferative effect on U937 cells evaluated by MTT in vitro methods and was much more efficient than that of Gleevec, which is known as a molecule targeting chemotherapeutical agent. The substitution of 2-phospholenes at C-3 and C-4 position with methyl groups as well as 4-bromo substituent suggests a good anti-proliferative effect.

Full text of DOI:10.1016/j.bmcl.2010.01.061

Bioorganic & Medicinal Chemistry Letters 20 (2010) 5943–5946
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Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Preparation and characterization of novel
4-bromo-3,4-dimethyl-1-phenyl-2-phospholene 1-oxide and the analogous
phosphorus heterocycles or phospha sugars
Manabu Yamada a, Mitsuji Yamashita a, , Takuya Suyama a, Junko Yamashita a, Kazuhide Asai a,
⇑
Taishi Niimi a, Nobuhisa Ozaki a, Michio Fujie a,b, Kasthuraiah Maddali a, Satoki Nakamura b,
Kazunori Ohnishi b
a Graduate School of Science and Technology, Shizuoka University, Naka-ku, Hamamatsu 432-8561, Japan
b Department of Internal Medicine III, Hamamatsu University School of Medicine, Kita-ku, Hamamatsu 431-3192, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
4-Bromo-3,4-dimethyl-1-phenyl-2-phospholene 1-oxide (3c) was first synthesized from 3,4-dimethyl-1-
phenyl-2-phospholene 1-oxide (2c) by a bromo-radical substitution reaction occurred at C-4 position by
N-bromosuccinimide and 2,20-azobisisobutyronitrile. The novel phospha sugar analogue 3c exerted high
anti-proliferative effect on U937 cells evaluated by MTT in vitro methods and was much more efficient
than that of GleevecÒ, which is known as a molecule targeting chemotherapeutical agent. The substitu-
tion of 2-phospholenes at C-3 and C-4 position with methyl groups as well as 4-bromo substituent sug-
gests a good anti-proliferative effect.
Received 26 November 2009
Revised 31 December 2009
Accepted 13 January 2010
Available online 21 January 2010
Keywords:
Phosphorus heterocycles
Phospha sugars
Tumors
Ó 2010 Elsevier Ltd. All rights reserved.
Leukemia cells
MTT in vitro method
Phospha sugars have a phosphorus atom instead of the oxygen
atom in the hemiacetal ring of the normal sugars and they con-
struct one new category of the pseudo sugars.1–4 Well known typ-
ical pseudo sugars are carba-, aza-, and thia-sugars,5–7 with a
carbon, a nitrogen, and a sulfur atom instead of the oxygen atom,
respectively, in the hemiacetal ring of the normal sugars, which
are known to exist in nature and are also prepared by synthetic su-
gar chemistry. Many of them exert important biological activities,
therefore, a lot of studies on them have actively been performed.
Recently bioactivities of phospha sugars were found, especially,
deoxybromophospha sugars exert important biological activities
and have quite efficient anti-cancer activities for leukemia cells.8
Since then researches on synthesis and evaluation of phospha sug-
ars are rapidly progressing to develop new type of molecular tar-
geting chemotherapeutic anti-tumor agents and the structure
and activity relationship and the mechanism of the action against
tumor cells are gradually been elucidated.9,10
alkyl group(s) and/or 4-position by a bromo group as well as no
substituents on the position, and the evaluation of the biological
activity for leukemia cells by using 3-(4,5-dimethyl-2-thiazolyl)-
2,5-diphenyltetrazolium bromide (MTT) in vitro methods.
Synthesis of 1-phenyl-2-phospholene 1-oxides 2a–c,11,12 being
used as the precursor of a pentofuranose-type phospha sugar,
was carried out by applying a known [4+2] cycloaddition reaction
(McCormac reaction)13 of 1,3-dienes 1a–c with phenylphospho-
nous dichloride and the successive hydrolysis reaction (Scheme
1). On the other hand, synthesis of 3-ethyl-1-phenyl-2-phospho-
lene 1-oxide (2d) was carried out by an alkylation reaction of the
methyl group of 3-methyl-1-phenyl-2-phospholene 1-oxide (2b)
by using n-buthyllithium and iodomethane (Scheme 2).
Ph
Ph
Cl
O
P
P
In this paper, we will deal with the research on novel phospha
sugar analogues or 2-phospholene derivatives 2 and 4-bromo-2-
phospholene derivatives 3 which were prepared from 2-phospho-
lene derivatives, being substituted on the 3- and/or 4-positions by
PhPCl2
H2O
R2
R1
1a-c
R2
R1
2a-c
R2
R1
Scheme 1. Preparation of 1-phenyl-2-phospholene 1-oxides 2a–c: (a) R1 = R2 = H;
⇑
Corresponding author.
E-mail address: tcmyama@ipc.shizuoka.ac.jp (M. Yamashita).
(b) R1 = CH3, R2 = H; (c) R1 = R2 = CH3.
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.01.061

5944
M. Yamada et al. / Bioorg. Med. Chem. Lett. 20 (2010) 5943–5946
stitution reaction with N-bromosuccinimide (NBS) and 2,20-azobis-
Ph
O
Ph
O
1.nBuLi
2.CH3 I
P
isobutyronitrile (AIBN) (Scheme 3). The allylic radical bromination
was occurred by the radical species, produced by thermal decom-
position of 2,20-azobisisobutyronitrile and NBS, which attacked at
the C-4 position of 2-phospholene 1-oxides 2. These results are
summarized in Table 1.14
P
CH3
C2H5
2b
2d
The prepared 2-phospholene 1-oxides 2 and 4-bromo-2-phos-
pholene 1-oxides 3 were bio-assayed by MTT in vitro method
against leukemia cells of U937 cell lines for the first time. The re-
sults against U937 cells are shown in Figures 1 and 2. U937 cells
of leukemia cell lines were incubated with 2-phospholene 1-oxides
2 or 4-bromo-2-phospholene 1-oxides 3 at the indicated concen-
Scheme 2. Preparation of 3-ethyl-1-phenyl-2-phospholene 1-oxide (2d).
Ph
O
Ph
O
1.NBS
2.AlBN
P
P
R2
trations (0–1000 lM) at 37 °C for 48 h. The cell proliferation or
R2
R1
R1
Br
inhibition was measured as the function of the absorbance at
560 nm visible light and evaluated by MTT in vitro assay.15
4-Bromo-3,4-dimethyl-2-phospholene 1-oxides 3c strongly
suppressed the cell proliferation of U937 cells in a dose-dependent
manner and the intensity of the absorbance at 560 nm decreased.
The decrease of the absorbance means the death of the leukemia
cells and clearly indicates that 3c possesses the growth inhibitory
effect on U937 cells.The results obtained by MTT in vitro evalua-
2a-d
3a-d
Scheme 3. Preparation of 4-bromo-2-phospholene 1-oxides 3a–d: (a) R1 = R2 = H;
(b) R1 = CH3, R2 = H; (c) R1 = R2 = CH3; (d) R1 = C2H5, R2 = H.
4-Bromo-2-phospholene derivatives 3a–d11,14 were prepared
from 2-phospholene compounds 2a–d by the bromo-radical sub-
Table 1
Preparation of 2-phospholene 1-oxides 2 and 4-bromo-2-phospholene 1-oxides 3
Entry
Substituent
R2
2-Phospholene 1-oxide 2
4-Bromo-2-phospholene 1-oxide 3
R1
Compound
Conditions
Yield (%)
Compound
Conditions
Yield (%)
1
2
3
4
H
H
H
CH3
H
2a
2b
2c
2d
CHCl3, rt, 3 weeks
CHCl3, rt, 2 weeks
CHCl3, rt, 2 weeks
THF, ꢀ78 °C, 24 h
22
68
30
34
3a
3b
3c
3d
CHCl3, reflux, 6 h
CHCl3, reflux, 6 h
CHCl3, reflux, 6 h
CHCl3, reflux, 6 h
26
65
31
44
CH3
CH3
C2H5
1.8
1.6
1.4
1.2
1.0
0.8
0.6
0.4
0.2
0
U937
2a
2b
2c
0
0.9
1.85
3.75
7.5
15.5
31
62.5
125
250
500
1000
Concentration (µM)
Figure 1. Evaluation by MTT in vitro assay for inhibition from cell proliferation of U937 cells by treatment with 2-phospholene 1-oxides 2a–2c at 37 °C for 48 h.
1.8
1.6
1.4
1.2
1.0
U937
0.8
3a
0.6
0.4
0.2
0
3b
3c
3d
0
0.9
1.85
3.75
7.5
15.5
31
62.5
125
250
500
1000
Concentration (µM)
Figure 2. Evaluation by MTT in vitro assay for inhibition from cell proliferation of U937 cells by treatment with 4-bromo-2-phospholene 1-oxides 3 at 37 °C for 48 h.

M. Yamada et al. / Bioorg. Med. Chem. Lett. 20 (2010) 5943–5946
5945
Table 2
9. Nakamura, S.; Yamashita, M.; Yokota, D.; Hirano, I.; Ono, T.; Fujie, M.; Shibata,
K.; Niimi, T.; Suyama, T.; Kasthuraiah, M.; Asai, K.; Yamashita, J.; Iguchi, Y.;
Ohnishi, K. Invest. New Drug 2009. doi:10.1007/s10637-009-9255-3.
Substituent effect of R1 and/or R2 at 3- and/or 4-positions of 1-phenyl-2-phospholene
1-oxides (2) on anti-proliferative effect by alkyl group of C-3 position
10. Yamada, M.; Asai, K.; Yamashita, J.; Suyama, T.; Niimi, T.; Kasthuraiah M.; Fujie,
M.; Nakamura, S.; Yamashita, M. Phosphorus, Sulfur, Silicon Relat Elem, in press.
11. General procedures and methods: TLC (Silica gel: Wako Chromato Sheet and/or
Merk Kieselgel 60; Eluent: CHCl3/MeOH = 20:1, in Rf value); Melting point
apparatus (Gallenkamp, in °C); MS (MALDI-TOF-MS: GL Science (Voyager-DE
Compound 2
Substituents at 3- and/or 4-position
IC50 (lM)
R1
R2
2a
2b
2c
H
H
>1000
900
350
Porimerix); Matrix: a-cyano-4-hydroxycinnamic acid, in m/z); IR (JASCO FT/IR
CH3
CH3
H
CH3
410 (KBr), in cmꢀ1); 1H NMR (JEOL JNM-AL300 (300 MHz); solvent: CDCl3, in d
(ppm)); and HPLC (JASCO and GL Science HPLC apparatuses, in tR) were used
for analyzing the products.
12. Preparation of 2-phospholenes 2: Synthesis of 3,4-dimethyl-1-phenyl-2-
phospholene 1-oxide (2c): 2,3-dimethyl-1,3-butadiene 10 ml (90 mmol;
1.0 equiv) and phenylphosphonous dichloride 16 ml (120 mmol; 1.30 equiv)
were mixed and reacted at room temperature for 2 weeks. The formed solid
3,4-dimethyl-1-phenyl-2-phospholenium dichloride was dissolved in
chloroform (200 ml) and hydrolyzed at 0 °C by addition of ice. The reaction
mixture was neutralized with sodium hydrogencarbonate and then filtered
and extracted with chloroform (20 ml ꢁ 3). The chloroform extract was
washed with water (50 ml), saturated sodium hydrogencarbonate solution
(50 ml), and saturated sodium chloride solution (50 ml). Drying over the
chloroform extract with anhydrous sodium sulfate followed by filtration,
evaporation of the solvent in vacuo, and distillation under the reduced
pressure afforded 3,4-dimethyl-1-phenyl-2-phospholene 1-oxide (2c, 5.6 g) in
30% yield. Bp: 130–132 °C/0.12 mmHg; Rf = 0.30 (CHCl3/MeOH = 20:1); 1H
NMR (CDCl3, 300 MHz) d = 1.24–1.37 (dd, 3H, C4–CH3), 1.80 (s, 3H, C3–CH3),
2.07–2.17 (ss, 1H, C4), 2.67–2.93 (m, 2H, C5), 5.89–5.97 (dd, 1H, C2), 7.48–7.75
(m, 5H, Ph) MALDI-TOF mass: m/z 205.7 (MH+, 50), 207.7 (MH+, 100); HPLC
(Wakosil 5SIL, CHCl3/MeOH = 20:1, flow rate 0.5 mL/min, k = 254 nm);
tR = 11.04 min.
Table 3
Substituent effect of R1 and/or R2 at 3- and 4-positions of 4-bromo-1-phenyl-2-
phospholene 1-oxides (3) on anti-proliferative effect by alkyl group of C-3 position
Compound 3
Substituents at 3- and/or 4-position
IC50 (lM)
R1
R2
3a
3b
3c
3d
H
CH3
H
H
>1000
87
CH3
CH2CH3
CH3
H
5.6
>1000
tion at the half of the absorbance intensity (IC50) against U937 cells
are shown in Tables 2 and 3. Tables 2 and 3 show that 4-bromo
substituent remarkably enhances the anti-leukemia activity.
Table 2 shows that 3,4-dimethyl-2-phospholene 1-oxide (2c) has
Similarly, 1-phenyl-2-phospholene 1-oxide (2a) and 3-methyl-1-phenyl-2-
phospholene 1-oxide (2b) were prepared.
1-Phenyl-2-phospholene 1-oxide (2a): Registry number: 703-03-7; Rf = 0.38
(CHCl3/MeOH = 20:1); bp 145–152 °C (0.08 mmHg); 1H NMR (CDCl3, 300 MHz)
d = 2.12–2.21 (m, 2H, C5), 2.78–2.94 (dd, 2H, C4), 6.25–6.36 (dt, 1H, C3), 7.05–
7.23 (dt, 1H, C2), 7.47–7.50 (m, 3H, m, p-Ph), 7.64–7.71 (m, 2H, o-Ph); MALDI-
TOF mass: m/z 179.6 (MH+, 100); HPLC (Wakosil 5SIL, CHCl3/MeOH = 20:1, flow
rate 0.5 mL/min, k = 254 nm); tR = 10.88 min.
3-Methyl-1-phenyl-2-phospholene 1-oxide (2b): Registry number: 707-61-9;
Rf = 0.32 (CHCl3/MeOH = 20:1); bp 148–161 °C (0.10 mmHg); 1H NMR (CDCl3,
300 MHz) d = 2.08 (s, 1H, –CH3), 2.17–2.29 (m, 2H, C5), 2.59–2.83 (dd, 2H, C4),
5.90–5.99 (dd, 1H, C2), 7.43–7.71 (m, 5H, Ph); MALDI-TOF mass m/e 193.7
(MH+, 100); HPLC (Wakosil 5SIL, CHCl3/MeOH = 20:1, flow rate 0.5 mL/min,
k = 254 nm); tR = 10.06 min.
3-Ethyl-1-phenyl-2-phospholene 1-oxide (2d): Under Ar atmosphere, to
tetrahydrofuran dehydrate (THF; 10 ml) solution of iodomethane (0.26 mL,
4.0 mmol, 2.0 equiv) was added n-hexane solution (1.6 mol/l; 2.55 ml,
4.0 mmol, 2.0 equiv) at ꢀ78 °C. After 2 h, 2b (384 mg, 2.0 mmol, 1.0 equiv) in
THF was added, and then the reaction mixture was stirred at ꢀ78 °C for 24 h.
After the completion of the reaction, the reaction mixture was allowed to stand
at room temperature and worked up by washing with 10% hydrochloric acid
(3 ml) and saturated ammonium chloride solution (3 ml), and then dried over
with anhydrous sodium sulfate and rotary evaporated. Purification of the
product through silica gel column (CHCl3/MeOH = 30:1–15:1) afforded 3-
ethyl-1-phenyl-2-phospholene 1-oxide (3; 205 mg, 1.00 mmol) in 50% yield.
a
higher activity than 3-methyl-2-phospholene 1-oxide (2b).
Table 3 shows that 3,4-dimethyl-4-bromo-2-phospholene 1-oxide
(3c) has the highest activity among 3 and 3-methyl derivative 3b is
much more active than 3a and 3d, whose R1 is H and C2H5, respec-
tively. The IC50 value of 3c was 5.6 lM. The observed anti-prolifer-
ative effect of 4-bromo-3,4-dimethyl-1-phenyl-2-phospholene
1-oxide 3c on U937 cells is much more efficient than that of
GleevecÒ (IC50 = 500
lM), which is clinically used as a molecule
targeting chemotherapeutical agent.9,10,16In conclusion, unsatu-
rated phospha sugars and unsaturated deoxybromo phospha
sugars or 2-phospholenes 2a–2c and 3a–3d were prepared. The
novel phospha sugar analogues 3b and 3c exerted anti-prolifera-
tive effect against U937 leukemia cells evaluated by MTT in vitro
methods. Especially 4-bromo-3,4-dimethyl-1-phenyl-2-phospho-
lene 1-oxide 3c, whose 3- and 4-positions were substituted with
methyl groups, possesses quite higher anti-cancer activity than
GleevecÒ (clinically being used against).
Formula:
C12H15OP; Exact Mass: 206.09; Registry number: 848484-65-1;
Rf = 0.29 (CHCl3/MeOH = 20:1); 1H NMR (CDCl3, 300 MHz) d = 2.13 (t, 3H, –
CH2CH3), 2.24–2.26 (m, 2H, C5), 2.38 (m, 2H, –CH2CH3), 2.71–2.86 (m, 2H, C4),
5.93–6.01 (dd, 1H, C2), 7.53–7.74 (m, 5H, Ph); MALDI-TOF mass: m/e 207 (MH+,
100).
Acknowledgements
13. Middlemas, E. D.; Quin, L. D. J. Org. Chem. 1979, 44, 2587.
The authors greatly acknowledge financial supports by the Sa-
tellite Shizuoka and the Headquarters of Japan Science and Tech-
nology Agency (JST) as well as Shizuoka Prefecture Industry
Creating Agency.
14. Preparation of 4-bromo-2-phospholenes 3: 4-Bromo-3,4-dimethyl-1-phenyl-2-
phospholene 1-oxide (3c): To a chloroform (3 ml) solution of 3,4-dimethyl-1-
phenyl-2-phospholene 1-oxide (2c; 206.1 mg, 1.00 mmol, 1.0 equiv) and N-
bromosuccinimide (NBS, 213.6 mg, 1.20 mmol, 1.2 equiv) was added dropwise
a chloroform (3 ml) solution of 2,20-azobisisobutyronitrile (AIBN, 24.6 mg,
0.15 mmol, 0.15 equiv) at 60 °C and the reaction mixture was refluxed for 6 h
under Ar atmosphere. The reaction mixture was neutralized with saturated
NaHCO3 aqueous solution (10 ml), washed with water (10 ml) and saturated
NaCl solution (10 ml), and dried over with anhydrous sodium sulfate. The
solvent of the filtrate was evaporated under a reduced pressure to give an oily
residual material. The residue was purified by column chromatography on
silica gel by using chloroform and methanol (20:1) as the eluent to give 4-
References and notes
1. Yamashita, M.; Yamada, M.; Nomoto, H.; Sugiura, M.; Oshikawa, T. Nippon
Kagaku Kaishi (J. Chem. Soc. Jpn.) 1987, 7, 1207.
2. Yamamoto, H.; Hosoyamada, C.; Kawamoto, H.; Inokawa, S.; Yamashita, M.;
Armour, M. A.; Nakashima, T. T. Carbohydrate Res. 1982, 102, 159.
3. Yamamoto, H.; Hanaya, T.; Kawamoto, H.; Inokawa, S.; Yamashita, M.; Armour,
M. A.; Nakashima, T. T. J. Org. Chem. 1985, 50, 3516.
4. Reddy, V. K.; Haritha, B.; Oshikawa, T.; Yamashita, M. Tetrahedron Lett. 2004, 45,
2851.
5. (a) Shan, M.; O’Doherty, G. A. Org. Lett. 2008, 10, 3381; (b) Sollogoub, M.; Sinay,
P. In Organic Chemistry of Sugars; Levy, D. E., Fugedi, P., Eds.; Taylor & Francis,
2006; p 349.
6. Alam, M. A.; Kumar, A.; Vankar, Y. D. Eur. J. Org. Chem. 2008, 4972.
7. Joseph, B.; Rollin, P. Phosphorus, Sulfur, Silicon 1993, 74, 467.
8. Ito, S.; Yamashita, M.; Niimi, T.; Fujie, M.; Reddy, V. K.; Totsuka, H.; Harutha, B.;
Maddali, K.; Nakamura, S.; Asai, K.; Suyama, T.; Yamashita, J.; Iguchi, Y.; Yu, G.;
Oshikawa, T. Heterocycl. Commun. 2009, 15, 23.
bromo-3,4-dimethyl-1-phenyl-2-phospholene
1-oxide
(3c;
0.089 g,
0.314 mmol) in 31% yield; TLC (Silica gel:Wako Chromato Sheet and/or Merk
Kieselgel 60; Eluent: CHCl3/MeOH = 20:1), Rf = 0.59; MS (MALDI-TOF-MS: GL
Science (Voyager-DE Porimerix); Matrix: a-Cyano-4-hydroxycinnamic acid (m/
z)), 285.3 (MꢀH+); 1H NMR (JEOL JNM-AL300 (300 MHz); solvent: CDCl3, d
(ppm)) = 1.73 (s, 3H, C3–CH3), 1.90–2.31 (m, 2H, C5), 2.21 (s, 3H, C4–CH3),
5.97–6.05 (dd, 1H, C-2), 7.48–7.79 (m, 5H, Ph).
Similar procedure for 3c afforded 3a, 3b, and 3d as follows:
4-Bromo-1-phenyl-2-phospholene 1-oxide (3a): Yield, 26 %; Rf = 0.49 (CHCl3/
MeOH = 20:1); MS (m/z), 285.3(MꢀH+); 1H NMR (CDCl3, 300 MHz),
d
(ppm) = 2.12–2.21(m, 2H, C5), 5.30–5.35 (t, 1H, C4), 6.26–6.37 (dt, 1H, C3),
7.06–7.24 (dt, 1H, C2) 7.48–7.71 (m, 5H, Ph).

5946
M. Yamada et al. / Bioorg. Med. Chem. Lett. 20 (2010) 5943–5946
4-Bromo-3-methyl-1-phenyl-2-phospholene 1-oxide (3b): Yield, 65 %; Rf = 0.42
dimethylsulfoxide (DMSO) (Sigma Chemical Company, St. Louis, MO, USA) as
the solvent, and were diluted into appropriate concentration with DMSO in
culture medium immediately before use. The final concentrations of 2 and 3 in
DMSO in all experiments were less than 0.010%, and all the treatment
conditions were compared with vehicle controls. The control experiments for
the evaluation were carried out by using DMSO, and the absorption change by
DMSO in the MTT method was not observed for U937 cells at 37 °C for 48 h.
16. (a) Capdeville, R.; Buchdunger, E.; Zimmermann, J.; Matter, A. Nat. Rev. Drug
Disc. 2002, 1, 493; (b) Arora, A.; Scholar, E. M. J. Pharmacol. Exp. Ther. 2005, 315,
971; (c) Melnikova, I.; Golden, J. Nat. Rev. Drug Disc. 2004, 3, 993; (d) Leonetti,
F.; Capaldi, C.; Carotti, A. Tetrahedron Lett. 2007, 48, 3455.
(CHCl3/MeOH = 20:1); MS (m/z), 284.0(MꢀH+); 1H NMR (CDCl3, 300 MHz), d
(ppm) = 2.22 (s, 1H, C3–CH3), 2.65–3.10 (m, 2H, C5), 4.96–5.17 (dd, 1H, C4),
5.91–6.38 (dd, 1H, C-2), 7.49–7.87(m, 5H, Ph).
4-Bromo-3-ethyl-1-phenyl-2-phospholene 1-oxide (3d): Yield, 44%; Rf = 0.25
(CHCl3/MeOH = 15:1); MS (m/z), 285.3(MꢀH+); 1H NMR (CDCl3, 300 MHz), d
(ppm) = 1.55–1.69 (t, 3H, C3–CH2CH3), 1.86–1.98 (m, 2H, C5), 2.14–2.18 (q, 2H,
C3–CH2CH3), 4.94 (s, 1H, C-4), 6.12–6.20 (dd, 1H, C-2), 7.74–7.99 (m, 5H, Ph).
15. MTT in vivo evaluation: Reagent and solvent for the in vitro MTT evaluation: 2-
Phospholene 1-oxides 2 and 4-bromo-2-phospholene 1-oxides 3, the reagents
being evaluated by the in vitro MTT method, were dissolved in