Tables at a glance

Table 1

Table 2

Figures at a glance

Comments

Figure 1

Comments

Figure 2

Comments

Figure 3

Comments

Figure 4

Comments

Figure 5

Comments

Figure 6

Comments

Figure 7

Comments

Figure 8

Comments

Figure 9

Comments

Figure 10

Comments

Figure 11

Comments

Figure 12

Table 1

Bioactive Compounds	Extracts
Bioactive Compounds	S. leucopyrus	V. negundo	A. indica	A. catechu	Kwath (aqueousextract)
Phenolics	+ve	+ve	+ve	+ve	+ve
Flavonoids	+ve	+ve	+ve	+ve	+ve
Tannins	+ve	+ve	+ve	+ve	+ve
Alkaloids	+ve	+ve	+ve	+ve	+ve
Terpenoids	+ve	+ve	+ve	+ve	+ve
Anthra-quinones	-ve	+ve	-ve	-ve	+ve
Saponins	-ve	+ve	+ve	+ve	+ve
Glycosides	-ve	+ve	+ve	+ve	+ve

Table 1 : Phytochemical Characterization: Qualitative methods

Table 1

Bioactive Compounds	Extracts
Bioactive Compounds	S. leucopyrus	V. negundo	A. indica	A. catechu	Formulation
Phenolics	226 µg/ml	61.76 µg/ml	158.16 µg/ml	1.87 µg/ml	34.908 µg/ml

Table 2 A : Test for Phenolics

Bioactive Compounds	Extracts
Bioactive Compounds	S. leucopyrus	V. negundo	A. indica	A. catechu	Formulation
Flavonoids	1.277 µg/ml	1.277 µg/ml	1.12 µg/ml	1.186 µg/ml	1.89 µg/ml

Table 2 B : Test for Flavonoids

Bioactive Compounds	Extracts
Bioactive Compounds	S. leucopyrus	V. negundo	A. indica	A. catechu	Formulation
Tannins	3.569 µg/ml	2.67 µg/ml	2.4249 µg/ml	0.146 µg/ml	1.27 µg/ml

Table 2 C : Test for Tannins

Bioactive Compounds	Extracts
Bioactive Compounds	S. leucopyrus	V. negundo	A. indica	A. catechu	Formulation
Alkaloids	17.2 µg/mg	21.67 µg/mg	18.35 µg/mg	11.2 µg/mg	17.1 µg/mg

Table 2 D : Test for Alkaloids

Bioactive Compounds	Extracts
Bioactive Compounds	S. leucopyrus	V. negundo	A. indica	A. catechu	Formulation
Terpenoids	0.976 mg	0.91 mg	0.879 mg	0.907 mg	mg

Table 2 E : Test for Alkaloids

Table 2 : Phytochemical Characterization: Quantitative methods

FIGURE 1

Figure 1: Antioxidant analysis by DPPH assay of oil and its extracts. The antioxidant activity of chloroform extract seems to be equivalent to NAC {N-acetyl cysteine, followed by aqueous extract, hexanoic extract and ethanoic extract showing nearly same potency, then petroleum ether extract is followed by our formulation, least in methanolic extract. The phytochemicals showed the highest antioxidant potency in semi-polar to non-polar solvents and the sesame oil is thereby used as a carrier, targeting the generation of oxidative stress at wound site.

FIGURE 2

Figure 2: GCMS Report

FIGURE 3

Figure 3: Cell proliferation assay (Dose Dependent) in Normal cell line. Cells were treated with varied concentration of oil. *p< 0.05

FIGURE 4

Figure 4: Cell proliferation assay (Dose Dependent) in Hyperglycemic cell line. Cells were treated with varied concentration of oil.*p< 0.05

FIGURE 5

Figure 5: Time dependent cell proliferation assay in Normal cell- 30µl (Orange) and hyperglycemic cell were treated with 0.5µl (Grey) and 1µl (Yellow). #p< 0.1, *p< 0.05

FIGURE 6

Figure 6: ROS generation in m5s cell. Cells were treated with various doses of Formulation for 2 hr and analysed by confocal and fluorescence microscopy. Representative images for (A) is Control and it gives green color fluorescence (B) Cells treated with 30µl oil. Cells with intracellular ROS are seen in less fluorescent. Control green colour compare to control. Intracellular ROS seen in more fluorescent compare to control. The green fluorescence represents stressful condition that remains almost similar in case of Control while in case of cells treated with oil, because of the potent antioxidant property, the ROS are scavenged. Hence the number of cell under stress is significantly less.

FIGURE 7

Figure 7: ROS generation in hyperglycaemic cells was (A) Control it gives green color fluorescence (B) and (C) Cells were treated with formulation (0.5µl, 1µl) showed less green fluorescence. (The green fluorescence represent stressful condition that remain almost similar in control while in case of cells treated with oil, because of the potent antioxidant property, the ROS are scavenged. Hence the number of cells under stress is significantly less

FIGURE 8

Figure 8: FACS performed in normal cells. A. The concentration of live cells in control is 95.03%, the cells undergoing pro-apoptosis are 3.73%. B. Cells treated with formulation are 94.7% showing similar result, the pro-apoptosis cell decrease to 3.17%

FIGURE 9

Figure 9: FACS performed in hyperglycaemic cells. (A) The concentration of live cells in control is 61.3%, cells treated with 0.5 µl (B) & 1 µl (C) formulation are 75.93% and 71.47% respectively. Under control condition the cells undergoing pro-apoptosis are 4.07%, while in 0.5 µl & 1 µl formulation treated cell decreases to 1.97% & 3.23% respectively

FIGURE 10

Figure 10: Phagocytosis activity in green fluorescence showed (A) control shows green fluorescence representing the phagocytic activity by macrophage. (B) and (C) Macrophage were treated with formulation showed significant phagocytic activity at 0.5µl and 1µl concentration

FIGURE 11

Figure 11: Cell migration assay (A) At 0 hr control, no migration was observed, (B) At 18hr control, cell migration elongation of dendrites in fibroblast and migration of border cells are indicative of invasiveness. (C) Formulation treated samples showed the significant migration of cells was observed

FIGURE 12

Figure 12: Cell Migration Assay in hyperglycaemic Cell (A) At 0 hr control, no migration was observed, (B) At 18hr control, cell migration elongation of dendrites in fibroblast and migration of border cells are indicative of invasiveness. (C) and (D) When treated with different concentrations of formulation i.e. 0.5µl and 1µl, gap filling started via cell migration