Synthesis of novel and stable 5-aminolevulinic acid derivatives for the efficient synthesis of 5-aminolevulinic acid based prodrugs

Article abstract of DOI:10.1055/s-2007-990825

The efficient synthesis of two stable, easily handled precursors of 5-aminolevulinate is reported. These precursors are stable to the chemical transformation needed for the synthesis of bioconjugates and can be readily deprotected in good yields. Georg Th

Full text of DOI:10.1055/s-2007-990825

PAPER
3731
Synthesis of Novel and Stable 5-Aminolevulinic Acid Derivatives for the
Efficient Synthesis of 5-Aminolevulinic Acid Based Prodrugs
Synthesis of
Nov
a
el and Stab
m
le 5-Aminolevulinic
a
A
cid Deri
k
vatives rishnan Vallinayagam,a Hugo Bertschy,b Yann Berger,c Virginie Wenger,a Reinhard Neier*a
a
Institut de Chimie, Université de Neuchâtel, Rue Emile-Argand, 11, PO 158, 2009 Neuchatel, Switzerland
Fax +41(32)7182511; E-mail: reinhard.neier@unine.ch
b
c
c/o F. Hoffmann-La Roche AG, Grenzacherstrasse 124, 4070 Basel, Switzerland
c/o Laboratoire cantonal contrôle denrées alimentaires, Les Croisettes, Case postale 68, 1066 Epalinges, Switzerland
Received 14 August 2007
Dedicated to Professor Wolf-Dietrich Woggon in honor of his 65th birthday
report the synthesis of an activated, stable, and storable
form of 5-aminolevulinic acid that can be utilized for its
efficient derivatization.
Abstract: The efficient synthesis of two stable, easily handled pre-
cursors of 5-aminolevulinate is reported. These precursors are sta-
ble to the chemical transformation needed for the synthesis of
bioconjugates and can be readily deprotected in good yields.
During our efforts to synthesize pseudopeptides contain-
Key words: amino acid, activated ester, protecting group, prodrug, ing 5-aminolevulinic acid, we tested several different pro-
tecting groups, such as the Fmoc or the Boc group.5 The
porphyrin
chemistry needed for deprotection of the Fmoc group was
not compatible with the reactivity of 5-aminolevulinic
acid and its derivatives in our hands. Boc-protected 5-
aminolevulinic acid (Boc-ALA) is easy to cleave without
affecting ester/amide functionalities. The Boc protection
renders 5-aminolevulinic acid stable, easily handled, and
assists purification.
Using 5-aminolevulinic acid (ALA) and its derivatives in
photodynamic therapy (PDT) as natural precursors of the
photosensitizer has gained increasing interest for the treat-
ment of different cancers.1 Successful clinical trials for tu-
mor-associated diseases such as actinic keratosis, basal
cell carcinoma, Bowen’s disease, and psoriasis have been
reported.2 5-Aminolevulinic acid–photodynamic therapy
involves administration of 5-aminolevulinic acid, or suit-
able derivatives thereof, which biosynthetically generate
protoporphyrin IX (PpIX), a powerful photosensitizer. On
application of light, protoporphyrin IX produces singlet
oxygen, which then destroys tumor and tumor-associated
cells. 5-Aminolevulinic acid is noninvasive and clears
rapidly from the body. 5-Aminolevulinic acid induced
protoporphyrin IX formation has been used not only in
cancer therapeutics but also in the detection of cancer
cells.3
Herein we report the efficient synthesis of Boc-ALA and
various activated esters of Boc-ALA suitable for the syn-
thesis of many different bioconjugates and other 5-ami-
nolevulinic acid derivatives. We synthesized Boc-ALA in
four steps from levulinic acid, an inexpensive and widely
available starting material6 in 20% overall yield
(Scheme 1).
O
O
O
a
b
OH
N3
O
Br
O
O
d
O
O
O
1
3
2
O
5-Aminolevulinic acid, an unusual amino acid, possesses
high water solubility and corresponding low lipid solubil-
ity and thereby has limited ability to cross biological
membranes. Various 5-aminolevulinic acid esters have
been synthesized to obtain more lipophilic prodrugs.4 5-
Aminolevulinic acid hydrochloride (ALA·HCl) is used as
the starting material for the synthesis of the corresponding
5-aminolevulinic acid prodrugs in most published reports.
5-Aminolevulinic acid hydrochloride is polar, difficult to
purify, and highly hygroscopic; it is difficult to handle and
forms side products at neutral pH, mostly due to reactions
between the oxo and the amino function. 5-Aminolevulin-
ic acid chloride has been used previously to derivatize 5-
aminolevulinic acid; it is unstable and must be generated
in situ and notoriously forms lactam-type impurities. We
c
BocHN
O
BocHN
OH
O
O
4
5
O
O
e
N3
O
N3
OH
O
O
6
3
Scheme 1 Reagents and conditions: (a) Br2, MeOH, r.t., overnight
then reflux, 1.5 h, 45%; (b) NaN3, THF–H2O, r.t., 1 h, 95%; (c) H2,
Pd/C, Boc2O, r.t., 24 h, 70%; (d) PLE, pH 8, 4 d, 64%; (e) PLE, pH 8,
10 h, 87%.
Levulinic acid (1) was brominated to give methyl 5-bro-
molevulinate (2) as the major regioisomer.7 Then, methyl
5-bromolevulinate (2) was converted into methyl 5-azi-
dolevulinate (3) by an SN2 reaction using sodium azide.
The reduction of the azide and in situ protection of the
amino group with the Boc group was initially attempted
by applying a Staudinger procedure, but this gave only
SYNTHESIS 2007, No. 23, pp 3731–3735
x
x.
x
x
.
2
0
0
7
Advanced online publication: 10.10.2007
DOI: 10.1055/s-2007-990825; Art ID: Z19107SS
© Georg Thieme Verlag Stuttgart · New York

3732
R. Vallinayagam et al.
PAPER
poor yields of 4.8a Hydrogenation of the azido group of tone oxime, respectively. Pentafluorophenyl 5-azidole-
methyl 5-azidolevulinate (3) using hydrogen and palladi- vulinate (10) was synthesized from 5-azidolevulinic acid
um on carbon in the presence of di-tert-butyl dicarbonate (6) in almost quantitative yield. These activated esters are
afforded cleanly methyl 5-(tert-butoxycarbonylamino)le- commonly used for the esterification of biomolecules
vulinate (4) in a satisfactory 70% yield.8b Using other such as monosaccharides.11
‘Boc’ reagents such as 2-[(tert-butoxycarbonyloxy)imi-
Thus, we have described the efficient and simple synthe-
sis of two stable, easily handled 5-aminolevulinic acid
no]-2-phenylacetonitrile (Boc-ON),9a tert-butyl amino-
carbonate,9b or tert-butyl carbonate (Boc2O) in the
precursors 5 and 6. Both precursors can be conveniently
presence of 4-(dimethylamino)pyridine did not improve
transformed into activated derivatives 7–10 that can be
used in place of 5-aminolevulinic acid hydrochloride for
the synthesis of 5-aminolevulinic acid prodrugs avoiding
most of the practical problems associated with the use of
the yield of 4. Hydrolysis of methyl 5-(tert-butoxycarbo-
nylamino)levulinate (4) was, in our hands, best performed
in the presence of the enzyme pig liver esterase (PLE) to
provide 5-(tert-butoxycarbonylamino)levulinic acid (5-
this sensitive compound.
Boc-ALA, 5) in 64% yield as a stable, crystalline solid.
Using our procedure 5-Boc-ALA 5 can be made in an eas-
ily handled free-flowing form. 5-Boc-ALA 5 can be
stored at room temperature for months without any signif-
icant decomposition, unlike ALA, which must be stored in
All moisture-sensitive reactions were carried out under argon or N2
using oven-dried glassware. All reagents were of the highest com-
mercial quality available if not specifically mentioned. Pig liver es-
terase (PLE, 24 units/mg) was purchased from Sigma-Aldrich.
the freezer to prevent its degradation.
Solvents were freshly distilled prior to use. Flash chromatography
(FC): SDS silica gel A C.C. Chromagel (35–70 mm); under positive
pressure, 0.5–0.9 bar. TLC: precoated silica gel thin-layer sheets 60
F 254 from Merck, detection by UV, basic KMnO4 soln or/and van-
illin soln. Refractive index (nD): Carl Zeiss; IR spectra: Perkin Elm-
er FT-IR Spectrum One version B; with a resolution of 2 cm–1.
Solids and oils were analyzed as KBr pellets, liquids as films be-
tween KBr plates. NMR Spectra: Bruker Avance-400 (400 and 100
MHz) or Varian Gemini XL-2000 (200 and 50 MHz), at r.t., if not
specified; Reference was TMS or the solvent peak as reference. The
sign denotes the calculated average value of J in cases where the
measured values varied up to 0.4 Hz. HETCOR (short range),
COSY and DEPT 135 were systematically measured for all com-
pounds to allow the attribution of all signals. MS: ESI and APCI:
Finnigan LCQ; the postulated formula of the peaks are indicated in
parenthesis; HRMS: Bruker FTMS 4.7T BioAPEXII measured at
University of Fribourg (Switzerland) by Mr. F. Nydegger.
Methyl 5-azidolevulinate (3) can also be hydrolyzed using
the same conditions to provide crystalline 5-azidolevulin-
ic acid (6), a masked form of ALA that can also be used
for the synthesis of various ALA prodrugs.
Starting from 5-Boc-ALA 5 we efficiently synthesized
various activated esters of 5-(tert-butoxycarbonylami-
no)levulinic acid suitable for further transformations
(Scheme 2). Pentafluorophenyl 5-(tert-butoxycarbonyl-
amino)levulinate (7), trichloroethyl 5-(tert-butoxycarbon-
ylamino)levulinate (8), and 5-(tert-butoxycarbonyl-
amino)levulinic acid acetone oxime ester (9) were synthe-
sized in good to excellent yields by coupling10 5-Boc-
ALA 5 with pentafluorophenol, trichloroethanol, and ace-
O
F
Methyl 5-Bromolevulinate (2)
a
BocHN
O
F
F
F
To a soln of levulinic acid (1, 100 g, 861 mmol, 1.0 equiv) in MeOH
(860 mL) at 0 °C was added dropwise over 3 h Br2 (44 mL, 861
mmol, 1.0 equiv); the temperature did not exceed 30 °C. The mix-
ture was brought to r.t. and stirred overnight and then the colorless
soln was refluxed for 90 min. MeOH was evaporated and the yellow
residue was dissolved in CH2Cl2 (400 mL). The organic phase was
washed with H2O (400 mL) followed by further washing with sat.
NaHCO3 (200 mL) and then with H2O (3 × 200 mL). The CH2Cl2
soln was dried (MgSO4), filtered through filter paper, and evaporat-
ed. The yellow residue (156 g) was subjected to fractional distilla-
tion using a Widmer column (40 cm) under reduced pressure (0.04
mbar) to give 2 (81 g, 45%); 98% purity (1H NMR); bp ~80 °C/
0.045 mbar; Rf = 0.50 (hexane–EtOAc, 1:1).
O
7
F
O
O
BocHN
OH
b
BocHN
O
CCl3
O
8
O
5
O
c
BocHN
O
N
nD20 1.4820.
O
9
IR (film): 3000 (m), 2953 (m), 2849 (w), 1734 (s), 1722 (s), 1439
(s), 1411 (s), 1359 (s), 1322 (m), 1207 (s), 1177 (s), 1079 (m), 1025
(m), 988 (m), 862 (w), 768 (vw), 694 (w), 595 (vw), 496 (vw), 464
(w) cm–1.
O
O
F
d
N3
OH
N3
O
F
F
O
O
F
10
6
1H NMR (200 MHz, CDCl3, 298 K): d = 3.95 (s, 2 H, H5), 3.67 (s,
F
3
3 H, CO2CH3), 2.95 (t, 3J3,2 = 6.4 Hz, 2 H, H3), 2.64 (t, J2,3 = 6.4
Scheme 2 Reagents and conditions: (a) pentafluorophenol, DCC,
CH2Cl2, r.t., overnight, 98%; (b) Cl3CCH2OH, EDC, DMAP, CH2Cl2,
r.t., overnight, 85%; (c) Me2C=NOH, EDC, DMAP, CH2Cl2, r.t.,
overnight, 84%; (d) pentafluorophenol, DCC, CH2Cl2, r.t., overnight,
98%.
Hz, 2 H, H2).
13C NMR (50 MHz, CDCl3, 298 K): d = 201.3 (C4), 173.4 (C1),
52.6 (CO2CH3), 35.1 (C3), 34.7 (C2), 28.8 (C5).
MS (ESI+): m/z = 231.1 [M(79Br) + Na]+, 233.0 [M(81Br) + Na]+.
Synthesis 2007, No. 23, 3731–3735 © Thieme Stuttgart · New York

PAPER
Synthesis of Novel and Stable 5-Aminolevulinic Acid Derivatives
3733
Methyl 5-Azidolevulinate (3)
1H NMR (400 MHz, CDCl3, 298 K): d = 11.30 (s, 1 H, COOH), 5.35
(br s, 1 H, NH), 4.03 (d, 3J5,NH = 4.8 Hz, 2 H, H5), 2.69–2.62 (m, 4
H, H2, H3), 1.40 [s, 9 H, C(CH3)3].
13C NMR (100 MHz, CDCl3, 298 K): d = 204.5 (C4), 177.4 (C1),
156.0 [C(O)t-Bu], 80.2 [C(CH3)3], 50.3 (C5), 34.2 (C3), 28.4
[C(CH3)3], 27.7 (C2).
To a soln of NaN3 (17.68 g, 272 mmol, 2 equiv) in deionized H2O
(68 mL) at 0 °C was added dropwise a soln of 2 (28.43 g. 136 mmol,
1 equiv) in THF (70 mL) . The yellow-colored mixture was brought
to r.t. and stirred for 1 h. The resulting two phases were separated.
The aqueous phase was extracted with EtOAc (3 × 200 mL). The
organic phases were combined and washed with H2O (2 × 200 mL),
dried (MgSO4), filtered, and evaporated to give 3 (22.05 g, 95%) as
a yellow oil; Rf = 0.50, 0.33 (UV254 active, vanillin) (hexane–
EtOAc, 2:1).
MS (APCI–): m/z (%) = 230.1 (95, [M – H]–).
5-Azidolevulinic Acid (6)
To a 0.1 M phosphate buffer soln (500 mL) was added, while stir-
ring the mixture, 3 (22.05 g, 128.8 mmol 1 equiv). Then pig liver
esterase (PLE) (300 mg, 24 units per mg) was added. The pH of the
mixture was maintained at pH 8 adding 5 M NaOH. The reaction
was followed by TLC (hexane–EtOAc, 1:1) and after 10 h was ex-
tracted with EtOAc (3 × 200 mL). The aqueous phase was acidified
to pH 2 with 6 M HCl. The aqueous phase was extracted with
EtOAc (4 × 500 mL). The combined organic phases were washed
with sat. NaCl (500 mL) followed by deionized H2O (500 mL). The
organic phase was dried (MgSO4) and evaporated. The crude solid
was triturated with Et2O at 0 °C. The pale brown pure solid thus ob-
tained was filtered and dried to give pure white 6 (17.61 g, 87%); Rf
= 0.35 (EtOAc), 0.91 (CH2Cl2–MeOH, 4:1) (UV254 active,
KMnO4).
IR (film): 2956 (m), 2914 (m), 2850 (w), 2543 (vw), 2202 (m), 2106
(vs), 1732 (vs), 1619 (w), 1438 (s), 1417 (s), 1361 (s), 1280 (s),
1210 (s),1172 (s), 1096 (s), 1028 (m), 1008 (m), 988 (m), 921 (m),
885 (w), 843 (w), 556 (m) cm–1.
1H NMR (200 MHz, CDCl3, 298 K): d = 4.03 (s, 2 H, H5), 3.69 (s,
3 H, CO2CH3), 2.76–2.72 (m, AA¢ part of a AA¢BB¢ system, 2 H,
H3), 2.70–2.66 (m, BB¢ part of a AA¢BB¢ system, 2 H, H2).
13C NMR (50 MHz, CDCl3, 298 K): d = 202.9 (C4), 173.0 (C1),
57.3 (C5; 51.9 (CO2CH3), 34.3 (C3), 27.4 (C2).
MS (ESI+): m/z = 194 [M + Na]+.
Methyl 5-(tert-Butoxycarbonylamino)levulinate (4)
A soln of 3 (23 g, 134.37 mmol, 1 equiv) in EtOAc (250 mL) was
charged in a hydrogenation autoclave, followed by Boc2O (29.63 g,
135.76 mmol, 1.01 equiv) and 10% Pd/C (5 g, 4.7 mmol, 0.035
equiv). The mixture was stirred mechanically during hydrogenation
at r.t. under 4.14 bar H2. After 24 h, the H2 pressure was released and
the mixture was filtered through Celite, which was washed with
EtOAc (3 × 25 mL); the filtrate was evaporated to give a yellow
oily. This crude mixture was purified by flash chromatography (gra-
dient, CH2Cl2–EtOAc 95:5 to CH2Cl2–EtOAc 85:15) to give pure 4
(23.05 g, 70%) as a yellow oil; Rf = 0.25–0.30 (UV254 active,
KMnO4) (CH2Cl2–EtOAc, 9:1).
IR (KBr): 3500–2500 (br s), 2914 (s), 2221 (m), 2110 (vs), 1720
(vs), 1411 (s), 1399 (s), 1374 (s), 1344 (m), 1288 (s), 1262 (s), 1235
(s), 1224 (s), 1173 (m), 1088 (s), 1046 (m), 1005 (w), 927 (s), 887
(m), 828 (w), 689 (w), 648 (w), 633 (m), 557 (m), 526 (w) cm–1.
1H NMR (400 MHz, CDCl3, 298 K): d = 9.48 (s, 1 H, COOH), 4.02
(s, 2 H, H5), 2.77–2.69 (m, 4 H, H2, H3).
13C NMR (100 MHz, CDCl3, 298 K): d = 203.0 (C4), 178.4 (C1),
57.6 (C5), 34.3 (C3), 27.7 (C2).
MS (APCI+): m/z (%) = 156.8 (60, [M – H]+).
IR (film): 2979 (s), 2932 (s), 1735 (s), 1719 (s),1701 (s), 1697 (s),
1523 (s), 1518 (s), 1508 (s), 1500 (s), 1438 (s), 1412 (s), 1392 (s),
1252 (s), 1209 (s),1165 (s), 1098 (m), 1060 (m), 1024 (m), 983 (m),
737 (m) cm–1.
1H NMR (400 MHz, CDCl3, 298 K): d = 5.24 (br s, 1 H, NH), 4.03
(d, 3J5,NH = 3.6 Hz, 2 H, H5), 3.64 (s, 3 H, CO2CH3), 2.71–2.68 (m,
2 H, H3), 2.63–2.59 (m, 2 H, H2), 1.40 [s, 9 H, C(CH3)3].
13C NMR (100 MHz, CDCl3, 298 K): d = 204.5 (C4), 173.0 (C1),
155.7 [C(O)t-Bu], 79.9 [C(CH3)3], 52.0 (CO2CH3), 50.3 (C5), 34.4
(C3), 28.4 [C(CH3)3], 27.6 (C2).
Pentafluorophenyl 5-(tert-Butoxycarbonylamino)levulinate (7)
To a soln of pentafluorophenol (2.7 g, 14.67 mmol, 1.1 equiv) in
CH2Cl2 (50 mL) at 0 °C, was added 5 (3 g, 12.97 mmol, 1 equiv)
followed by DCC (3 g, 14.54 mmol, 1.12 equiv). The reaction was
brought to r.t. and kept overnight. The mixture was filtered over
Celite and the solvent was evaporated. The thick yellow oily mass
obtained was triturated with hexane to remove excess pentafluo-
rophenol and the white solid obtained was filtered. The white solid
was dried to give pure 7 (5.1 g, 98%); Rf = 0.33 (CH2Cl2–EtOAc,
95:5) (UV254 active; KMnO4).
IR (KBr): 3009 (w), 2982 (w), 2948 (w), 2920 (w), 1798 (m), 1725
(s), 1694 (vs), 1656 (w), 1626 (w), 1527 (vs), 1518 (vs), 1468 (w),
1446 (w), 1426 (w), 1401 (w), 1392 (w), 1370 (m), 1359 (w), 1328
(w), 1271 (s), 1253 (m), 1235 (w), 1170 (m), 1146 (m), 1111 (s),
1042 (m), 1031 (m), 1007 (s), 989 (s), 977 (m), 895 (w), 864 (w),
609 (w), 556 (w) cm–1.
1H NMR (400 MHz, CDCl3, 298 K): d = 5.25 (br s, 1 H, NH), 4.06
(d, 3J5,NH = 4.9 Hz, 2 H, H5), 2.99 (t, 3J2,3 = 6.5 Hz, 2 H, H3), 2.87
(t, 3J3,2 = 6.5 Hz, 2 H, H2), 1.43 [s, 9 H, C(CH3)3].
MS (APCI+): m/z (%) = 269.1 (15, [M + Na]+).
5-(tert-Butoxycarbonylamino)levulinic Acid (5)
To a 0.1 M phosphate buffer soln (500 mL) of pH 8 was added with
stirring 4 (22.85 g, 93.2 mmol, 1 equiv). Then pig liver esterase
(PLE) (300 mg, 24 units per mg) was added. The pH of the mixture
was maintained at pH 8 by addition of 5 M NaOH. The reaction was
followed by TLC (CH2Cl2–EtOAc, 90:10). After 4 d the mixture
was extracted with EtOAc (3 × 200 mL), the aqueous phase was
acidified to pH 2 with 6 M HCl, and extracted with EtOAc (4 × 500
mL). The organic phases were combined and washed with sat. NaCl
(500 mL) followed by deionized water (500 mL). The organic phase
was dried (MgSO4) and evaporated. A yellow crude solid product
was obtained by trituration with Et2O at 0 °C to give pure white 5
(13.7 g, 64%); Rf = 0.47 (KMnO4, tailing) (EtOAc).
13C NMR (100 MHz, CDCl3, 298 K): d = 203.6 (C4), 168.8 (C1),
1
155.8 [C(O)t-Bu], 142.4 (d m, JC–F = 248 Hz, C2Pfp, C3Pfp, or
1
C4Pfp), 139.9 (d m, JC–F = 244 Hz, C2Pfp, C3Pfp, or C4Pfp), 137.5
1
(d m, JC–F = 240 Hz, C2Pfp, C3Pfp, or C4Pfp), 125.0 (C1Pfp), 80.2
[C(CH3)3], 50.3 (C5), 34.2 (C3), 28.4 [C(CH3)3], 27.0 (C2).
19F NMR (188 MHz, CDCl3, 298 K): d = –152.9 (m, 2 F, C2Pfp
C2¢Pfp), –158.2 (t, 3JF–F ≈ 21.0 Hz, 1 F, (C4Pfp), –162.6 (m, 2 F, C3Pfp
C3¢Pfp).
,
,
IR (KBr): 3379 (s), 3500–2500 (br s), 2986 (m), 2919 (m), 1707
(vs), 1686 (vs), 1506 (s), 1429 (s), 1407 (m), 1392 (m), 1383 (m),
1371 (m), 1363 (m), 1349 (m), 1295 (w), 1278 (m), 1240 (s), 1209
(s),1170 (s), 1098 (w), 1065 (m), 1039 (vw), 1015 (m), 943 (m), 894
(m), 872 (m), 803 (w), 787 (w), 759 (w), 741 (w), 653 (w), 575 (m)
cm–1.
MS (ESI+): m/z (%) = 419.9 [M + Na]+.
Synthesis 2007, No. 23, 3731–3735 © Thieme Stuttgart · New York

3734
R. Vallinayagam et al.
PAPER
2,2,2-Trichloroethyl 5-(tert-Butoxycarbonylamino)levulinate
(8)
1H NMR (400 MHz, CDCl3, 298 K): d = 4.06 (s, 2 H, H5), 3.06 (t,
3J2,3 = 6.5 Hz, 2 H, H3), 2.94 (t, 3J3,2 = 6.5 Hz, 2 H, H2).
To a soln of 2,2,2-trichloroethanol (1.267 g, 8.48 mmol, 2 equiv) in
CH2Cl2 (80 mL) at 0 °C, DMAP (0.528 g, 4.32 mmol, 1 equiv) was
added followed by EDC (0.912 g, 4.75 mmol, 1.1 equiv) and 5 (1 g,
4.32 mmol, 1 equiv). The mixture was brought to r.t. and kept over-
night. The solvent was evaporated and the mixture was dissolved in
EtOAc (100 mL). The organic phase was extracted with H2O (200
mL).The organic phase was washed with cold 1 M citric acid (170
mL), followed by sat. NaHCO3 (200 mL) and sat. NaCl (200 mL).
The organic phase was dried (MgSO4) and the solvent was evapo-
rated. The remaining crude product was purified by trituration with
hexane to remove excess alcohol and the white solid obtained was
filtered. The white solid was dried to yield 8 (1.33 g, 85%); Rf = 0.17
(UV254 active; KMnO4) (hexane–EtOAc, 7:3).
13C NMR (100 MHz, CDCl3, 298 K): d = 202.3 (C4), 168.9 (C1),
1
141.5 (d m, JC–F = 252 Hz, C2Pfp, C3Pfp, or C4Pfp), 140.0 (d m,
1JC–F = 254 Hz, C2Pfp, C3Pfp, or C4Pfp), 138.2 (d m, 1JC–F = 250 Hz,
C2Pfp, C3Pfp, or C4Pfp), 125.0 (C1Pfp), 57.8 (C5), 27.0 (C2).
MS (APCI–): m/z (%) = 322.2 (95, [M – H]–).
Acknowledgment
This research was supported by the Swiss National Science Found-
ation (Funding number 3152A0-102184) and the University of
Neuchatel. NMR and mass spectra were measured on instruments at
the Universities of Neuchatel and Fribourg acquired with the assi-
stance of grants from the Swiss National Science Foundation. We
acknowledge the contribution from graduate students C. Kopp, J.
Weber, and P. Gurba for this work.
IR (KBr): 3000 (w), 2981 (w), 2932 (w), 1759 (vs), 1721 (vs), 1688
(vs), 1528 (vs), 1477 (w), 1452 (w), 1408 (m), 1387 (m), 1369 (s),
1357 (m), 1333 (w), 1283 (vs), 1252 (m), 1195 (m), 1175 (s), 1156
(vs), 1134 (vs), 1084 (w), 1070 (w), 1043 (m), 1029 (w), 1006 (vw),
968 (w), 904 (w), 867 (vw), 831 (m), 797 (m), 716 (s), 681 (w), 654
(w), 571 (w) cm–1.
References
1H NMR (400 MHz, CDCl3, 298 K): d = 5.24 (br s, 1 H, NH), 4.70
(s, 2 H, CH2CCl3), 4.04 (d, 3J5,NH = 5.0 Hz, 2 H, H5), 2.77 (s, 4 H,
H2, H3), 1.41 [s, 9 H, C(CH3)3].
13C NMR (100 MHz, CDCl3, 298 K): d = 204.0 (C4), 170.9 (C1),
155.7 [C(O)t-Bu], 94.8 (CH2CCl3), 80.0 [C(CH3)3], 74.2
(CH2CCl3), 50.3 (C5), 34.2 (C3), 28.4 [C(CH3)3], 27.5 (C2).
(1) (a) Fukuda, H.; Casas, A.; Batlle, A. Int. J. Biochem. Cell
Biol. 2005, 37, 272. (b) Dickson, E. F. G.; Kennedy, J. C.;
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M.; Wagnieres, G.; Juillerat-Jeanneret, L.; Van den Bergh,
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Jeanneret, L. J. Med. Chem. 2000, 43, 4738. (b) Berger, Y.;
Ingrassia, L.; Neier, R.; Juillerat-Jeanneret, L. Bioorg. Med.
Chem. 2003, 11, 1343. (c) Greppi, A. Ph.D. Thesis;
University of Neuchâtel: Switzerland, 1999.
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MS (APCI+): m/z (%) = 263.9 (99, [M(35Cl) – Boc + H]+), 265.9
(32, [M(37Cl) – Boc + H]+).
5-(tert-Butoxycarbonylamino)levulinic Acid Acetone Oxime Es-
ter (9)
To a soln of acetone oxime (0.643 g, 8.8 mmol, 1 equiv) in CH2Cl2
(85 mL) at 0 °C, DMAP (1.18 g, 9.68 mmol, 1.1 equiv) was added
followed by EDC (2.05 g, 10.69 mmol, 1.2 equiv) and 5 (2.03 g,
8.80 mmol, 1 equiv). The mixture was brought to r.t. and was kept
overnight. The solvent was evaporated and the residue was purified
by flash chromatography (Et2O) to give 9 (2.13 g, 84%); Rf = 0.19
(UV254 active; KMnO4) (Et2O).
IR (KBr): 3005 (m), 2978 (s), 2922 (m), 1771 (vs), 1705 (vs), 1651
(m), 1519 (vs), 1457 (m), 1440 (m), 1411 (s), 1397 (s), 1367 (vs),
1287 (vs), 1251 (s), 1164 (vs), 1127 (vs), 1070 (s), 1054 (m), 1944
(m), 1027 (m), 935 (m), 952 (w), 935 (w), 885 (vs), 784 (m), 771
(w), 747 (w), 588 (m), 577 (w), 548 (w), 464 (w) cm–1.
1H NMR (400 MHz, CDCl3, 298 K): d = 5.24 (br s, 1 H, NH), 4.03
(d, 3J5,NH = 5.0 Hz, 2 H, H5), 2.76–2.69 (m, 4 H, H2, H3), 1.99 [s, 3
H, N=C(CH3)2], 1.96 [s, 3 H, N=C(CH3)2], 1.45 [s, 9 H, C(CH3)3].
13C NMR (100 MHz, CDCl3, 298 K): d = 204.4 (C4), 170.4 (C1),
164.2 (CO2CH3), 155.7 [C(O)t-Bu], 80.0 [C(CH3)3], 50.3 (C5), 34.2
(C3), 28.4 [C(CH3)3], 26.6 (C2), 21.9, 17.0 [N=C(CH3)2].
Pentafluorophenyl 5-Azidolevulinate (10)
To a soln of pentafluorophenol (2.2 g, 11.9 mmol, 1.1 equiv) in
CH2Cl2 (40 mL) at 0 °C, was added 6 (1.72 g, 11 mmol, 1 equiv)
followed by DCC (2.5 g, 12.3 mmol, 1.12 equiv). The reaction was
brought to r.t. and kept overnight. The mixture was filtered over
Celite and the solvent was evaporated. The thick yellow oil obtained
was triturated with hexane to remove excess pentafluorophenol and
the white solid was filtered to give pure 10 (3.47 g, 98%) as a white
solid. Rf = 0.36 (UV254 active; KMnO4) (CH2Cl2–EtOAc, 95:5).
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1994, 24, 2557.
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Synthesis of Novel and Stable 5-Aminolevulinic Acid Derivatives
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Synthesis 2007, No. 23, 3731–3735 © Thieme Stuttgart · New York