Curcuminoid-Derived 3,5-Bis (Styryl) Isoxazoles - Mechanochemical Synthesis and Antioxidant Activity
Curcuminoid-Derived 3,5-Bis (Styryl) Isoxazoles - Mechanochemical Synthesis and Antioxidant Activity
Curcuminoid-Derived 3,5-Bis (Styryl) Isoxazoles - Mechanochemical Synthesis and Antioxidant Activity
c Indian Academy of Sciences.
DOI 10.1007/s12039-016-1119-8
1. Introduction 2. Experimental
2.3b Ferric ion reducing antioxidant power (FRAP) and control, respectively. At and Aot are the absorbance
assay: Freshly prepared FRAP reagent was used values of the sample and control after incubation time.
for the analysis. It was obtained by mixing acetate
buffer (300 mM each of acetic acid and sodium
acetate), tripyridyltriazine (10 mM in 40 mM aqueous
hydrochloric acid solution) and FeCl3 .6H2 O (20 mM) 3. Results and Discussion
aqueous solutions in the ratio 10:1:1. Methanolic solu-
tion of FeSO4 .6H2 O was prepared at different con- 3.1 Synthesis
centrations (0.05, 0.1, 0.25, 0.5, 0.75, 1.0, 1.5 and
2.0 mM). The solutions of bis(styryl)isoxazoles (4a- The precursor curcuminoids (3a-g) were obtained by
g) were also prepared in methanol (0.5 mM) and after microwave assisted synthetic protocol23 by the modi-
adding these (0.04 mL) to the FRAP reagent (3 mL), the fication of a conventional thermal method developed
reaction mixtures were incubated at 37◦ C for 30 min. earlier by our group.14 Sui et al., seemingly in the first
The absorbance was measured at 596 nm and the results report on the synthesis of an isoxazole derivative (2c) of
were expressed as micro molar Fe2+ equivalents. a curcuminoid, had reacted curcumin I (1) with hydrox-
ylamine hydrochloride in ethanol under reflux for
16 h.13 The methods subsequently reported for this reac-
2.3c β-Carotene bleaching assay: β-Carotene (0.2 mg), tion could be seen to generally take 6–40 h for the com-
linoleic acid (0.02 mL) and Tween 20 (0.18 mL) were pletion under conventional heating.5 9 In continuation
mixed with chloroform (0.5 mL) and the solvent was to our work on mechanochemical synthesis of curcumin
removed by evaporation. The residue obtained was derivatives,19 we have now reacted curcuminoids (3a-
mixed with distilled water (50 mL) and the result- g) with hydroxylamine hydrochloride in presence of
ing emulsion (4 mL aliquots each) was added to catalytic amount of acetic acid. The room temperature
3,5-bis(styryl)isoxazoles (4a-g) in methanol (0.2 mL, mechanical grinding in an agate mortar for about 1–
1 mmol). Absorbance was measured at 470 nm, using a 2 minutes rapidly afforded the 3,5-bis(styryl)isoxazoles
mixture of the emulsion (4 mL) and methanol (0.2 mL) (4a-g) in powder form. We observed that the com-
as the control. The absorbance was again recorded after pounds obtained in better purity and yield than reported
incubation for 1 h at 50◦ C in the dark. The antioxidant earlier, as judged by TLC. The formation and identity of
activity (AA) was evaluated in terms of bleaching of the these isoxazoles were confirmed by physical and spec-
β-carotene using the formula AA = [1 − (Ao – At )/(Aoo – tral methods. It appears that the initially formed oxime
Aot )] ×100, where Ao and Aoo are the absorbance values derivatives proceeded to cyclize affording the isoxazole
measured at zero time of the incubation for test sample ring (scheme 1). Among the 3,5-bis(styryl)isoxazoles
O OH
H3CO OCH3
1
HO OH
N X
H3CO OCH3
O OH N O
(4a-g) now synthesized, five examples (4b, 4c, 4d, 4f Table 2. The FeSO4 equivalent of 3,5-bis(styryl)isoxazo-
and 4g) appear to be hitherto unreported. les (4a-g) in the presence of FRAP reagent and the Antioxi-
dant Activity (AA) % of 4a-g in β–carotene assay.
4a 1182 87.59
The antioxidant (AO) activities of the 3,5-bis(styryl) 4b 1195 90.91
isoxazoles (4a-g) were evaluated by 2,2-diphenylpicryl- 4c 997 76.99
hydrazyl (DPPH), ferric ion reducing antioxidant power 4d 883 69.52
(FRAP) and β-carotene assays. The AO activity by the 4e 592 65.31
DPPH assay is expressed in terms of EC50 values, cal- 4f 1036 80.23
4g 475 61.12
culated by plotting the percentage inhibition of DPPH
against various test concentrations (figure 1, table 1).
We have earlier shown that the isoxazole derivative of a methoxy group on the terminal aryl rings enhances the
curcumin I (2c) exhibits better AO capacity than the AO activity of curcuminoid derived isoxazoles.
parent curcumin I (1) in DPPH assay under similar The results of FRAP assay are expressed as micro-
experimental conditions;19 the corresponding EC50 val- molar Fe2+ equivalents (table 2); a higher value of fer-
ues for the percentage inhibition of DPPH being 8 ± rous sulphate equivalent being indicative of a better AO
0.011 μmol and 40 ± 0.06 μmol, respectively for (2c) capacity. Our earlier sttudies19 have shown that under
and (1). A comparison of this data with those obtained identical assay conditions, the curcumin I derived isox-
now for the 3,5-bis(styryl)isoxazoles (4a-g) shows that azole (2c) and curcumin I (1) showed Fe2+ equiva-
the isoxazoles (4a-g) are less active than the isoxa- lent of 1304 μmol and 1164 μmol, respectively. The
zole derivative of curcumin I (2c). However, two among present results based on FRAP assay is now seen to
these, 4a and 4b, are more active than curcumin I (1). follow a similar pattern as seen in the DPPH assay,
Thus, the presence of a hydroxy group juxtaposed with wherein the presence of a hydroxy group along with a
methoxy group on the terminal aryl rings is found to
100 be beneficial. A similar conclusion could now be drawn
4a based on the β-carotene bleaching assay of the 3,5-
80 4b bis(styryl)isoxazoles (4a-g) (table 2) and in the light of
our earlier study.19
% inhibition
4c
60
4d
4e 4. Conclusions
40
4f
We report the synthesis of curcuminoid-derived 3,5-
20 4g
bis(styryl)isoxazoles (4a-g) by mechanochemical grind-
ing of the respective curcuminoid and hydroxylamine
0 hydrochloride catalyzed by gl. acetic acid. The anti-
0 0.02 0.04 0.06 0.08 0.1 0.12
oxidant studies of 4a-g revealed that hydroxy and
Conentration (mmol)
methoxy groups present in the terminal aryl moieties
Figure 1. Percentage inhibition of DPPH (10−2 mM) at of 3,5-bis(styryl)isoxazoles improve their anti-oxidant
various concentrations of 3,5-bis(styryl)isoxazoles (4a-g). activity.
4a 23.91 ± 0.21 37.15 ± 0.06 49.98 ± 0.31 60.29 ± 0.18 79.08 ± 0.12 87.42 ± 0.16 28 ± 0.19
4b 27.12 ± 0.18 40.36 ± 0.11 54.25 ± 0.22 64.29 ± 0.37 82.13 ± 0.19 91.05 ± 0.16 20 ± 0.11
4c 18.28 ± 0.23 26.32 ± 0.09 33.74 ± 0.16 39.62 ± 0.22 47.64± 0.07 63.29 ± 0.34 80 ± 0.48
4d 16.27 ± 0.16 19.11 ± 0.23 27.18 ± 0.17 31.13 ± 0.25 40.77 ± 0.06 53.02 ± 0.14 96 ± 0.52
4e 15.03 ± 0.29 18.20 ± 0.18 24.09 ± 0.08 28.78 ± 0.24 35.43 ± 0.17 48.78 ± 0.09 102 ± 0.49
4f 19.26 ± 0.33 29.88 ± 0.17 38.56 ± 0.24 47.23 ± 0.29 55.64 ± 0.07 74.28 ± 0.13 60± 0.31
4g 13.26 ± 0.11 16.27 ± 0.21 21.78 ± 0.09 25.98 ± 0.12 33.06 ± 0.19 46.52 ± 0.17 108 ± 0.22
3,5-Bis(styryl)isoxazoles 1319