Arch. Pharm. Chem. Life Sci. 2011, 11, 358–365
Benzothieno[3,2-e][1,2,4]triazolo[4,3-c]pyrimidines
363
3-(2-Chloro-phenyl)-9,10,11,12-tetrahydro[1]
SLN formulation
Preparation of triazolothienopyrimidines loaded SLNs
benzothieno[3,2-e][1,2,4]triazolo[4,3-c]pyrimidine (3c)
Yield: 88%, m.p.: 161–1638C. IR (KBr) cmꢀ1: 3090, 2921, 2866,
1625, 1573, 1541; 1H-NMR (CDCl3) d: 1.97 (m, 4H, CH2), 2.94
(t, J ¼ 8 Hz, 2H, CH2), 3.23 (t, J ¼ 8 Hz, 2H, CH2), 7.40–8.06 (m,
4H, Ar-H), 9.23 (s, C5-H pyrimidine proton); 13C-NMR d: 22.4, 23.6,
25.6, 25.8, 115.7, 120.6, 129.5, 129.6, 129.9, 130.3, 135.8, 139.9,
150.1, 153.7, 163.2, 164.4, 166.3; mass m/z: 340.8 (Mþ). Anal. calcd.
for C17H13ClN4S: C, 59.91; H, 3.84; N, 16.44; S, 9.41. Found: C,
59.22; H, 3.56; N, 15.94; S, 9.24.
SLNs formulations were prepared from a warm o/w microemul-
sion technique [23]. The synthesized compound 3h was dispersed
in molten lipid (708C), warm aqueous solution of egg lecithin was
added to melted lipid-drug mixture (at 708C) in presence of the
co-surfactant Tween 80 under stirring. The warm microemulsion
was then added carefully drop wise into ice cold water (2–38C)
with continuous stirring (T25 basic Ultra Turrax, IKA, USA). The
ratio between the microemulsion and the dispersion medium
was about 1:10. The dispersion was subjected to ultrasonication
for a period of 10 min to form nanosuspension (Table 2).
3-(3-Benzyl)-9,10,11,12-tetrahydro[1]benzothieno-
[3,2-e][1,2,4]triazolo[4,3-c]pyrimidine (3d)
Yield: 78%, m.p.: 138–1408C. IR (KBr) cmꢀ1: 3092, 2924, 2868,
1628, 1573, 1541; 1H-NMR (CDCl3) d: 1.96 (m, 4H, CH2),
2.94(t, J ¼ 8 Hz, 2H, CH2), 3.24 (t, J ¼ 8 Hz, 2H, CH2), 3.82 (s,
2H, -CH2-), 7.32–7.45 (m, 5H, Ar-H), 9.19 (s, 1H, C5-H pyrimidine
proton); mass m/z: 320.1 (Mþ). Anal. calcd. for C18H16N4S: C, 67.47;
H, 5.03; N, 17.49; S, 10.01. Found: C, 67.20; H, 5.38; N, 17.98; S,9.90.
Particle size analysis and zeta potential
Size and zeta potential of molecule loaded SLN were measured by
Photon Correlation Spectroscopy (PCS) using zetasizer 3000 HSA
(Malvern, U.K.). Samples were diluted appropriately with the
aqueous phase of the formulation for the measurements and
the pH of diluted samples ranged from 6.8 to 7.0.
3-(Naphthalen-2-yl)-9,10,11,12-tetrahydro[1]
Entrapment efficiency
The entrapment efficiency of the synthesized molecule was
determined by measuring the concentration of free drug in
the dispersion medium [23]. The sample was centrifuged at
4000 rpm for 30 min. The amount of free drug was determined
in the clear supernatant by UV spectrophotometer using super-
natant of non-loaded nanoparticles as basic correction. The
amount of incorporated compound was determined by subtract-
ing amount of initial compound from the free compound.
The entrapment efficiency was calculated by the following
equation [24]:
benzothieno[3,2e][1,2,4]triazolo[4,3-c]pyrimidine (3e)
Yield: 78%, m.p.: 227–2298C. IR (KBr) cmꢀ1: 3064, 2927, 2857,
1619; 1H-NMR (300 MHz,CDCl3) d: 1.98 (m, 4H, CH2), 2.93
(t, J ¼ 8 Hz, 2H, CH2), 3.25 (t, J ¼ 8 Hz, 2H, CH2), 7.56–8.44 (m,
6H, Ar-H), 9.21 (d, J ¼ 9 Hz, 1H, C2 naphthalene), 9.27 (s, C5-H,
pyrimidine proton); mass m/z: 356.4 (Mþ). Anal. calcd.
for C21H16N4S; C, 70.76; H, 4.52; N, 15.72; S, 9.00. Found: C,
70.12; H, 4.20; N, 15.92; S, 9.04.
3-(3-Methyl-thiophen-2-yl)-9,10,11,12-tetrahydro[1]
ꢀ
ꢁ
benzothieno[3,2-e][1,2,4]triazolo[4,3-c]pyrimidine (3f)
Yield: 80%, m.p.: 202–2048C. IR (KBr) cmꢀ1: 3054, 2932, 2855,
1617. 1H-NMR (CDCl3) d: 1.98 (m, 4H, CH2), 2.36 (s, 3H, CH3), 2.93
(t, J ¼ 8 Hz, 2H, CH2), 3.25 (t, J ¼ 8 Hz, 2H, CH2), 7.18 (d, 1H,
J ¼ 6 Hz, thiophenic proton), 7.95 (d, 1H, J ¼ 6 Hz, thiophenic
proton), 9.11 (s, C5-H pyrimidine proton); mass m/z: 326.3 (Mþ).
Anal. calcd. for C16H14N4S2: C, 58.87; H, 4.32; N, 17.16; S, 19.65.
Found: C, 58.28; H, 4.10; N, 17.32; S, 19.82.
Winitial compoundꢀWfree compound
EEð%Þ ¼
ꢃ 100
Winitial compound
where Winitial compound is the weight of the initial compound
and Wfree compound the weight of the free compound.
Scanning electron microscopy (SEM)
The morphological examination of 3h-SLN3 was performed by
scanning electron microscopy (Joel JSM 840A, Japan). Cleaned
brass specimen studs were used for mounting the samples. Wet
solvent paint was applied on these studs and while the paint was
wet, the pellets were placed on each stud and allowed to dry. The
sample was observed by SEM.
3-(Pyren-1-yl)-9,10,11,12-tetrahydro[1]benzothieno-
[3,2-e][1,2,4]triazolo[4,3-c]pyrimidine (3g)
Yield: 63%, m.p.: 216–2188C. IR (KBr) cmꢀ1: 3065, 2928, 2857,
1
1619; H-NMR (CDCl3) d: 1.98 (m, 4H, CH2), 2.93 (t, J ¼ 8 Hz, 2H,
CH2), 3.25 (t, J ¼ 8 Hz, 2H, CH2), 7.71–8.04 (m, 9H, Ar-H), 9.13
(s, C5-H pyrimidine proton); mass m/z: 434.5 (Mþ). Anal. calcd.
for C27H22N4S: C, 74.63; H, 5.10; N, 12.89; S, 7.38. Found: C, 74.42;
H, 5.56; N, 12.08; S, 7.64.
Differential scanning calorimetry
Differential scanning calorimetry (DSC) analysis of the drug,
bulk lipid and nanoparticles was conducted using a differential
scanning calorimeter (DSC Q20 V24.4 Build 116, TA Instruments,
USA) set at a heating rate of 108C/min.
3-(3-Methyl-furan-2-yl)-9,10,11,12-tetrahydro[1]
benzothieno[3,2e][1,2,4]triazolo[4,3-c]pyrimidine (3h)
Yield: 88%, m.p.: 196–1988C. IR (KBr) cmꢀ1: 3065, 2928, 2856,
1620; 1H-NMR (CDCl3) d: 1.98 (m, 4H, CH2), 2.21 (s, 3H, CH3), 2.93
(t, J ¼ 8 Hz, 2H, CH2), 3.24 (t, J ¼ 8 Hz, 2H, CH2), 7.08 (d, J ¼ 6 Hz,
1H), 7.45 (d, J ¼ 6 Hz, 1H), 9.14 (s, C5-H, pyrimidine proton); mass
m/z: 310.3 (Mþ). Anal. calcd. for C16H14N4OS: C, 61.92; H, 4.55; N,
18.05; S, 10.33. Found: C, 61.62; H, 4.85; N, 17.95; S, 10.03.
Short-term stability study
The selected SLN formulation was stored at 408C, 75% RH, in a
stability chamber (Thermo Lab., Mumbai) for a period of one
month and average particle size and entrapment efficiency was
determined.
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