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4.3. Dimethyl 3,30-(7,12-bis(1-pentadecanoxyethyl)-3,8,13,17-
tetramethylporphyrin-2,18-diyl)dipropionate (3b)
MS (ESI, positive): m/z (%): 963.7; HRMS (ESI, positive) m/z Calcd
for C60H90N4O6 [M+H]+; 963.6939, found: 963.6939.
This compound was synthesized from compound 1 (590 mg,
1.0 mmol) and 1-pentadecanol (503 mg, 2.2 mmol) using the pro-
cedure described for 3a to give 3b (200 mg 18% yield) as a dark vio-
let solid: Rf = 0.8 (CH2Cl2/MeOH 20:1); 1H NMR (400 MHz, CDCl3,
TMS) d 10.61 (s, 1H), 10.59 (s, 1H), 10.10 (s, 1H), 10.07 (s, 1H),
6.10 (m, 2H), 4.40–4.44 (m, 4H), 3.71 (s, 3H), 3.70 (s, 3H), 3.69 (s,
3H), 3.68 (s, 3H), 3.67 (s, 3H), 3.65 (s, 3H), 3.64 (m, 4H), 3.27–
3.31 (m, 4H), 2.24–2.25 (d, J = 3.4 Hz, 6H), 1.07–1.80 (m, 50H),
0.84 (t, J = 7.0 Hz, 6H; CH3); 13C NMR (75 MHz, CDCl3, TMS) d
173.6 (2C; C@O), 140.4, 138.3, 137.3, 137.0 (16C; C pyrrole), 98.8,
98.5, 96.8, 96.1, 73.3 (2C; OCH), 69.6 (2C; OCH2), 51.7 (2C;
COOCH3), 37.0, 31.9 (2C; CH2COOMe), 30.3, 29.6, 29.5, 29.3 26.4,
25.5 (26C; –CH2–), 22.7 (2C; CH2CH2COOMe), 21.9 (2C; –CH2–),
14.1 (2C; CH3C pyrrole), 11.8 (2C; CH3C pyrrole) ppm; IR (NaCl
disc) 3311, 2923, 2853, 1739, 1457, 1166 cmꢀ1; MS (ESI, positive):
m/z (%): 1047; HRMS (ESI, positive) m/z Calcd for C66H102N4O6
[M+H]+; 1047.7878, found: 1047.7878.
4.6. 3,30-(7,12-Bis(1-pentadecanoxyethyl)-3,8,13,17-tetramethyl-
porphyrin-2,18-diyl)dipropionic acid (4b: PL-C15)
This compound was synthesized from compound 3b (105.0 mg,
0.1 mmol) and LiOHꢁH2O (9.23 mg, 0.22 mmol) using the proce-
dure described for 4a to give 4b (91.8 mg, 90% yield) as a dark vio-
let solid; Rf = 0.8 (CH2Cl2/MeOH 20:1); 1H NMR (400 MHz, CDCl3,
TMS) d 10.67 (br s, 2H), 10.11 (br s, 2H), 6.10 (m, 2H), 4.43 (m,
4H), 3.74 (s, 3H), 3.72 (s, 3H), 3.68 (s, 3H), 3.66 (s, 3H), 3.62 (m,
4H), 3.34 (m, 4H), 2.24 (m, 6H), 1.09–1.78 (m, 50H), 0.83–0.87
(m, 6H); 13C NMR (300 MHz, CDCl3, TMS) d 179.9, 179.6 (2C;
C@O), 140.8, 138.0, 136.7 (16C; C pyrrole), 99.2, 98.8, 97.2, 95.7
(4C; porphine), 73.3 (2C; OCH), 69.6 (2C; OCH2), 37.5 (2C, CH3CH),
31.9 (2C; CH2COOMe), 30.3, 29.7, 29.5, 29.6, 29.3, 26.4, 25.5 (26C; –
CH2–), 22.7 (2C; CH2CH2COOMe), 22.0 (2C; –CH2–), 14.1 (2C; CH3C
pyrrole), 11.7, 11.6 (2C; CH3C pyrrole) ppm; IR (NaCl disc): 3311,
2923, 2853, 1739, 1455, 1165 cmꢀ1; MS (ESI, positive): m/z
(%):1019; HRMS (ESI, positive) m/z Calcd for C64H98N4O6 [M+H]+;
1019.7565, found: 1019.7568.
4.4. Dimethyl 3,30-(7,12-bis(1-heptadecanoxyethyl)-3,8,13,17-
tetramethylporphyrin- 2,18-diyl)dipropionate (3c)
4.7. 3,30-(7,12-Bis(1-heptadecanoxyethyl)-3,8,13,17-tetramethyl-
porphyrin-2,18-diyl)dipropionic acid (4c: PL-C17)
This compound was synthesized from compound 1 (590 mg,
1.0 mmol) and 1-heptadecanol alcohol (564 mg, 2.2 mmol) using
the procedure described for 3a to give 3c (210 mg 21% yield) as a
dark violet solid: Rf = 0.8 (CH2Cl2/MeOH 20:1); 1H NMR
(400 MHz, CDCl3, TMS) d10.62 (s, 1H), 10.60 (s, 1H), 10.11 (s, 1H),
10.07 (s, 1H), 6.08 (m, 2H), 4.04–4.44 (m, 4H), 3.74 (s, 3H), 3.72
(s, 3H), 3.70 (s, 3H), 3.69 (s, 3H), 3.68 (s, 3H), 3.67 (s, 3H), 3.65
(m, 4H), 3.28–3.31 (m, 4H), 2.24–2.26 (d, J = 6.4 Hz, 6H), 1.08–
1.77 (m, 58H), 0.86 (t, J = 6.8 Hz, 6H; CH3); 13C NMR (75 MHz,
CDCl3, TMS) d 173.6 (2C; C@O), 140.4, 138.3, 137.1, 136.7 (16C; C
pyrrole), 98.82, 98.54, 97.73, 96.04, 73.3 (2C; OCH), 69.6 (2C;
OCH2), 51.7 (2C; COOCH3), 36.9, 31.9 (2C; CH2COOMe), 30.3, 29.7,
29.6, 29.6, 29.5, 29.5, 29.3, 26.4, 25.5 (30C; –CH2–), 22.7 (2C;
CH2CH2COOMe), 21.9 (2C; –CH2–), 14.1 (2C; CH3C pyrrole), 12.5
(2C; CH3C pyrrole) ppm; IR (NaCl disc): 3311, 2924, 2852, 1739,
1435, 1166 cmꢀ1; MS (ESI, positive): m/z (%): 1103; HRMS (ESI,
positive) m/z Calcd for C70H110N4O6 [M+H]+; 1103.8504, found:
1103.8504.
This compound was synthesized from compound 3c (110 mg,
0.1 mmol) and LiOHꢁH2O (9.23 mg, 0.22 mmol) using the proce-
dure described for 4a to give 4c (100 mg, 93% yield) as a dark violet
solid; Rf = 0.8 (CH2Cl2/MeOH 20:1); 1H NMR (400 MHz, CDCl3, TMS)
d 10.67 (br s, 1H), 10.65 (br s, 1H), 10.12 (br s, 1H), 10.10 (br s, 1H),
6.12 (m, 2H), 4.45–4.51 (m, 4H), 3.77 (s, 3H), 3.75 (s, 3H), 3.73 (s,
3H), 3.70 (s, 3H), 3.64 (m, 4H), 3.41–3.42 (m, 4H), 2.24–2.27 (m,
6H), 1.10–1.84 (m, 58H), 0.85–0.90 (m, 6H); 13C NMR (300 MHz,
CDCl3, TMS) d 180.1 (2C; C@O), 140.6, 137.9, 137.0, 136.5 (16C; C
pyrrole), 99.1, 98.8, 96.9, 95.5 (4C; porphine), 73.3 (2C; OCH),
69.6 (2C; OCH2), 37.6 (2C, CH3CH), 31.9 (2C; CH2COOMe), 30.3,
30.2, 29.7, 29.6, 29.5, 29.3, 26.5, 26.4, 25.5, 25.4 (30C; –CH2–),
22.7 (2C; CH2CH2COOMe), 22.0 (2C; –CH2–), 14.1 (2C; CH3C pyr-
role), 11.8 (2C; CH3C pyrrole) ppm; IR (NaCl disc): 3311, 2923,
2852, 1709, 1463, 1099 cmꢀ1; MS (ESI, positive): m/z (%):1075;
HRMS (ESI, positive) m/z Calcd for
C
70H106N4O6 [M+H]+;
1075.8191, found: 1075.8183.
4.5. 3,30-(7,12-Bis(1-tridecanoxyethyl)-3,8,13,17-tetramethyl-
porphyrin-2,18-diyl)dipropionic acid (4a: PL-C13)
4.8. Preparation and characterization of PPIX–lipid micelles
Compound 3a (99.1 mg, 0.1 mmol) was dissolved in a THF/
MeOH/H2O mixture (15 mL, 1:1:1). LiOHꢁH2O (9.23 mg,
0.22 mmol) was added and the reaction mixture was stirred for
6 h at room temperature. The reaction mixture was filtered and
the volume of the filtrate was reduced under vacuum (5 mL). HCl
(1 N, 2 mL) was slowly added to this solution with stirring to pre-
cipitate the dark violet solid, which was filtered off and then
washed with water (5 mL) to give 4a (86.8 mg, 90% yield) as a dark
violet solid. Rf = 0.8 (CH2Cl2/MeOH 20:1); 1H NMR (400 MHz,
CDCl3, 25 °C, TMS) d 10.67 (br s, 1H), 10.65 (br s, 1H), 10.11 (br s,
2H), 6.08 (m, 2H), 4.45 (m, 4H), 3.73 (s, 3H), 3.69 (s, 3H), 3.67 (s,
3H) 3.65 (s, 3H), 3.63 (m, 4H), 3.36–3.37 (m, 4H), 2.23–2.24 (d,
J = 2.4 Hz, 6H), 1.08–1.80 (m, 42H), 0.83 (t, J = 7.0 Hz, 6H); 13C
NMR (300 MHz, CDCl3, TMS) d 179.6 (2C; C@O), 145.2, 140.8,
140.1 (16C; C pyrrole), 99.2, 98.8, 97.2 (4C; porphine), 73.3 (2C,
OCH) 69.6 (2C, OCH2), 37.5 (2C, CH3CH), 31.8 (2C; CH2COOMe),
30.3, 29.7, 29.5, 29.3, 26.4, 25.5 (24C; –CH2–), 22.6 (2C;
CH2CH2COO), 14.1 (2C, CH3C pyrrole), 11.7 ppm (2C; CH3C pyr-
To a THF solution (3 mL) of PPIX–lipids (PL-C13, PL-C15, or PL-
C17; 0.01 mmol) was added aqueous lithium hydroxide solution
(0.02 M, 1 mL) and the mixture was stirred for 1 h. After THF was
removed under the reduced pressure, the resulting lithium salts
(5a–c) of PPIX lipids were dissolved in water (1 mL) to form
micelles. The micelle solution was filtrated three times through a
Minisart filter with 0.45
lm pore size (Sigma Aldrich, 16555 K,
Minisart Filter, 0.45 m pore size). Micelle size was measured with
l
an electrophoretic light scattering spectrophotometer (Nano-ZS,
Sysmex, Japan). UV/vis spectra were measured with a Shimadzu
2450 PC spectrophotometer over the 300–700 nm wavelength
range.
4.9. Cytotoxicity of PPIX–lipid micelles
HeLa cells were used for the cell viability assay. The cells
(5 ꢂ 103 cells per well of a 96-well plate) were incubated at 37 °C
for 72 h in RPMI-1640 medium (100 lL) containing the PPIX–lipid
role); IR (NaCl disc): 3309, 2923, 2853, 1713, 1456, 1100 cmꢀ1
;
micelles at various concentrations. After the incubation, 30-(4,5-
dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT,