S1
Supporting Information
O-Benzoyl Pyridine Aldoxime and Amidoxime Derivatives:
Novel Efficient DNA Photo-Cleavage Agents
Paraskevi Karamtzioti,a Asterios Papastergiou,a John G. Stefanakis,b Alexandros E. Koumbis,b
Ioanna Anastasiou,a Maria. Koffa,a Konstantina C. Fylaktakidoua,*
a Laboratory of Organic, Bioorganic and Natural Product Chemistry, Molecular Biology and
Genetics Department, 68100 Alexandroupolis, Greeceb Laboratory of Organic Chemistry, Chemistry Department, Aristotle University of Thessaloniki,
54124 Thessaloniki, Greece
E-mail: [email protected]; tel: ++30-25510-30663; fax: ++30-25510-30613
Electronic Supplementary Material (ESI) for Medicinal Chemistry Communications.This journal is © The Royal Society of Chemistry 2015
S2
Table of contents
1H and 13C NMR spectra of 2 S3-S41H and 13C NMR spectra of 3 S5-S61H and 13C NMR spectra of 4 S7-S81H and 13C NMR spectra of 6 S9-S101H and 13C NMR spectra of 7 S11-S121H and 13C NMR spectra of 8 S13-S141H and 13C NMR spectra of 9 S15-S161H and 13C NMR spectra of 14 S17-S181H and 13C NMR spectra of 15 S19-S201H and 13C NMR spectra of 16 S21-S221H and 13C NMR spectra of 17 S23-S24
Photochemistry S25
Irradiation of N΄-((4-nitrobenzoyl)oxy)picolinimidamide (15) in MeOH/H2O 9:1 S25
Irradiation of N΄-((4-nitrobenzoyl)oxy)picolinimidamide (15) in benzene S27
Gel electrophoresis of plasmid with or without N΄-((4-nitrobenzoyl)oxy)
isonicotinimidamide (17) at various pHs S30
UV-Vis spectra of compounds 4, 8, 14-17 in DMSO S30
S3
S4
S5
S6
S7
S8
S9
S10
S11
S12
S13
S14
S15
S16
S17
S18
S19
S20
S21
S22
S23
S24
S25
PhotochemistryIrradiation of derivative 15 under various conditions at 312 nm.
1. Irradiation of N΄-((4-nitrobenzoyl)oxy)picolinimidamide (15) in MeOH/H2O 9:1.
N΄-((4-nitrobenzoyl)oxy)picolinimidamide 15 (13 mg) was placed into a quartz tube containing
a mixture of MeOH/H2O 9:1 (2 mL). The resulting heterogeneous mixture was degassed for 1 h
and then irradiated at 312 nm for 5 h. The floating solid was removed by filtration to give 9 mg
of unreacted starting material. Samples of the solid and the filtrate were analyzed with LC-MS.
N
N
NH2
O
O
NO2
hv
MeOH/H2O
N
NH2
NH
N
NH2
O
N
NH2
NOH
O2N
O
OH
+ + +
O2N
O O
NO2
+
N
NH2
N
N
NH2
NO
O2N
O
O
+ +
O2N
O+
N
NH2
NO N
NH2N+
15
Scheme 1. Proposed structures for the products obtained by the irradiation N΄-((4-
nitrobenzoyl)oxy)picolinimidamide 15 in MeOH/H2O 9:1. The assignment is based on the
fragments observed by the analysis of the LC-MS.
Solid
UV detection MS detection
0.0 1.0 2.0 3.0 4.0 5.0 6.0 7.0 min
0.0
0.5
1.0
1.5
2.0
2.5
3.0
mAU (x100)254nm,4nm (1.00)
0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 5.0 5.5 6.0 6.5 7.0 7.5 8.0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
1.0
1.1(x10,000,000)
2:TIC1:TIC
Time 4.9 min
Positive: 287 [M+H]+, 309 [M+Na]+, 341 [M+Na+MeOH]+.
50 100 150 200 250 300 350 400 450 500 550 m/z0.0
1.0
2.0
3.0
4.0
5.0
6.0
7.0Inten. (x100,000)
309
287
341
449120
224 403246 489 593378152 552 5674171928067 178
N
N
NH2
O
O
NO2
Chemical Formula: C13H10N4O4Exact Mass: 286,070
S26
Filtrate
UV detection MS detection
0.0 1.0 2.0 3.0 4.0 5.0 6.0 7.0 8.0 min
-0.1
0.0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
1.0
1.1
mAU (x100)254nm,4nm (1.00)
0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 5.0 5.5 6.0 6.5 7.0 7.5 8.0 8.5 9.0
1.0
2.0
3.0
4.0
5.0
6.0
7.0
8.0
(x1,000,000)
2:TIC1:TIC
Time 3.1 min
50 100 150 200 250 300 350 400 450 500 550 m/z0.00
0.25
0.50
0.75
1.00
Inten. (x1,000,000)
122
105137
235198 278 326 462169 34062 358 417 486
Positive: 122 [M+H]+.
Time 4.9 min
50 100 150 200 250 300 350 400 450 500 550 m/z0.0
0.5
1.0
1.5
2.0
2.5
3.0Inten. (x100,000)
579279
333102
138
192
123 224 528380 438
65 311181 581441400266 525477
Positive: 123 [M1+H]+, 138 [M2+H]+, 279 [M4+Na]+, 333 [M3+H+MeOH]+.
50 100 150 200 250 300 350 400 450 500 550 m/z0.00
0.25
0.50
0.75
1.00
1.25
1.50
1.75
2.00
2.25Inten. (x100,000)
279
579333
528309
250 380
477224 441400 524 587438120 152 170102 19158
Positive: 279 [M1+Na]+, 287 [M2+H]+, 309 [M2+Na]+, 333 [M3+H+MeOH]+.
N
NH2
NH
Chemical Formula: C6H7N3Exact Mass: 121,0640
N
NH2
O
Chemical Formula: C6H6N2OExact Mass: 122,048
M1N
NH2
NOH
Chemical Formula: C6H7N3OExact Mass: 137,0589
M2
N
N
NH2
O
O
NO2
Chemical Formula: C13H10N4O4Exact Mass: 286,0702
M2
O2N
O O
NO2
Chemical Formula: C14H8N2O6Exact Mass: 300,0382
M3N
NH2
NO N
NH2N
Chemical Formula: C12H12N6OExact Mass: 256,1073
M1
N
NH2
NO N
NH2N
Chemical Formula: C12H12N6OExact Mass: 256,1073
M4
O2N
O O
NO2
Chemical Formula: C14H8N2O6Exact Mass: 300,0382
M3
S27
Time 6.8 min
50 100 150 200 250 300 350 400 450 500 550 m/z0.0
0.5
1.0
1.5
2.0
2.5
3.0
3.5
4.0
Inten. (x100,000)
166
355
538522266198 300 431 455 556122 224 48337193 58540313863
Negative: 166 [M-H]-.
The starting material 15 is highly insoluble in the reaction solvent. From the integration
of the UV detection of the filtrate it seems that the N-O cleavage dominates, as fragment with
m/z 166 appears in high percentage. Amidine is consumed in side reactions.
2. Irradiation of N΄-((4-nitrobenzoyl)oxy)picolinimidamide (15) in benzene.
N΄-((4-nitrobenzoyl)oxy)picolinimidamide 15 (13 mg) was placed into a quartz tube containing
benzene (2 mL). The resulting heterogeneous mixture was degassed for 1 h, 1,4-cyclohexadiene
(10 eq) was added and then irradiated at 312 nm for 5 h. The floating solid was removed by
filtration to give 8 mg of unreacted starting material. Samples of the solid and the filtrate were
analyzed with LC-MS.
N
N
NH2
O
O
NO2
hv
1,4-cyclohexadiene,benzene
N
NH2
NH
N
NH2
O
N
NH2
NOH
O2N
O
OH
+ + + +
N
NH2
N
N
NH2
NO
O2N
O
O
+ +
O2N
O+
N
NH2
NO N
NH2NN
NH2
NNH2N
N +
15
Scheme 2. Proposed structures for the products obtained by the irradiation N΄-((4-
nitrobenzoyl)oxy)picolinimidamide 15 in benzene. The assignment is based on the fragments
observed by the analysis of the LC-MS.
O2N
O
OHChemical Formula: C7H5NO4
Exact Mass: 167,0219
S28
Solid
UV detection MS detection
0.0 1.0 2.0 3.0 4.0 5.0 6.0 min-0.25
0.00
0.25
0.50
0.75
1.00
1.25
1.50
mAU (x100)254nm,4nm (1.00)
0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0 4.5 5.0 5.5 6.0 6.5 7.0
1.0
2.0
3.0
4.0
5.0
6.0
7.0
8.0
9.0(x1,000,000)
2:TIC1:TIC
Time 4.9 min
Same as before for solid.
Filtrate
UV detection MS detection
0.0 2.5 5.0 7.5 10.0 min
-0.25
0.00
0.25
0.50
0.75
1.00
1.25
1.50
1.75
mAU (x100)254nm,4nm (1.00)
0.0 1.0 2.0 3.0 4.0 5.0 6.0 7.0 8.0 9.0 10.0 11.0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
1.0
(x10,000,000)
2:TIC1:TIC
Time 3.1 min
50 100 150 200 250 300 350 400 450 500 550 m/z0.0
1.0
2.0
3.0
4.0
5.0
Inten. (x100,000)
241122
105
154 259 275 346 361168 200 322 489418 537 58967 555399 517443
Positive: 122 [M1+H]+, 241 [M2+H]+.
Time 4.6 min
50 100 150 200 250 300 350 400 450 500 550 m/z0.00
0.25
0.50
0.75
1.00
Inten. (x100,000)
123
264202 296
180 505234
390317 360 53365 461102 576343145599
479438
Positive: 123 [M+H]+.
N
NH2
NH
Chemical Formula: C6H7N3Exact Mass: 121,0640
M1
N
NH2
O
Chemical Formula: C6H6N2OExact Mass: 122,0480
N
NH2
NNH2N
N
Chemical Formula: C12H12N6Exact Mass: 240,1123
M2
S29
Time 4.9 min
50 100 150 200 250 300 350 400 450 500 550 m/z0.0
0.5
1.0
1.5
2.0
2.5
3.0
3.5
4.0
Inten. (x100,000)
138
279
309
341 380224265 528105 557400 477188 587524424 44910467
Positive: 138 [M1+H]+, 279 [M2+Na]+, 309 [M3+Na]+.
Time 6.8 min
50 100 150 200 250 300 350 400 450 500 550 m/z0.0
0.5
1.0
1.5
2.0
2.5
3.0
3.5
4.0Inten. (x100,000)
166
355538522
198 327300266224 450 48337193 585407135122 57262
Negative: 166 [M-H]-.
The starting material 15 is highly insoluble in the reaction solvent. From the integration
of the UV detection of the filtrate it seems that the N-O cleavage dominates, as fragment with
m/z 166 appears in high percentage. Amidine is consumed in side reactions.
References for irradiation of oxime acyl compounds: a) J J.-P. Vermes and R. Beugelmans, Tetrahedron Lett., 1969, 10, 2091–2092; b) J. Lalevée, X. Allonas, J. P. Fouassier, H. Tachi, A. Izumitani, M. Shirai and M. Tsunooka, J. Photochem. Photobiol. A, Chem., 2002, 151, 27–37; c) R. Alonso, P. J. Campos, M. A. Rodríguez and D. Sampedro, J. Org. Chem., 2008, 73, 2234–2239.
N
N
NH2
O
O
NO2
Chemical Formula: C13H10N4O4Exact Mass: 286,0702
M3
N
NH2
NOH
Chemical Formula: C6H7N3OExact Mass: 137,059
M1
O2N
O
OHChemical Formula: C7H5NO4
Exact Mass: 167,022Molecular Weight: 167,119
N
NH2
NO N
NH2N
Chemical Formula: C12H12N6OExact Mass: 256,1073
M2
S30
Gel electrophoresis of plasmid with or without N΄-((4-nitrobenzoyl)oxy)
isonicotinimidamide (17) at various pHs
1 Plasmid + tris buffer pH 42 Plasmid + 17 (500 μΜ) – tris buffer pH 4 + UV3 Plasmid + tris buffer pH 54 Plasmid + 17 (500 μΜ) – tris buffer pH 5 + UV5 Plasmid + tris buffer pH 66 Plasmid + 17 (500 μΜ) – tris buffer pH 6 + UV7 Plasmid + tris buffer pH 78 Plasmid + 17 (500 μΜ) – tris buffer pH 7 + UV 9 Plasmid + tris buffer pH 810 Plasmid + 17 (500 μΜ) – tris buffer pH 8 + UV11 Plasmid + tris buffer pH 912 Plasmid + 17 (500 μΜ) – tris buffer pH 9 + UV13 Plasmid + tris buffer pH 1014 Plasmid + 17 (500 μΜ) – tris buffer pH 10 + UV
UV-Vis spectra of compounds 4, 8, 14-17 in DMSO