Background
Methods
Plant materials and reagents
Extraction, fractionation and isolation
Assay method of PTP1β inhibitory activity
Kinetics of PTP1β by active compounds
Extraction process
Experimental design for RSM
Total polyphenol content determination
Total anthocyanin content determination
Determination of extraction yield
Data analysis
Result
PTP1β inhibitory compounds from Zea mays L
Extract and fractions | Concentration (μg/mL) | Inhibition (%) | IC50b (μg/mL) |
---|---|---|---|
Suramina | 12.97 | 61.88 | |
6.49 | 40.62 | 7.51 | |
3.21 | 10.69 | ||
EtOH ext. | 100 | 43.99 | - |
n-Hex fr. | 100 | 25.54 | - |
CH2Cl2 fr. | 100 | 28.12 | - |
EtOAc fr. | 100 | 83.88 | |
50 | 57.23 | 26.12 | |
10 | 44.77 | ||
n-BuOH fr. | 100 | 70.89 | |
50 | 45.09 | 58.20 | |
10 | 26.87 | - | |
Water fr. | 100 | 47.50 | - |
Isolation of the compounds from active EtOAc fraction
Compounds | Concentration (μg/mL) | Inhibition (%) | IC50b (μg/ml) | IC50 (μM) |
---|---|---|---|---|
Suramina | 5 | 64.32 | 2.76 | |
2.5 | 46.46 | 3.94 | ||
1 | 10.69 | |||
Protochtechuic acid (1) | 100 | <50.0 | - | - |
Vanillic acid (2) | 100 | <50.0 | - | - |
2,4,6-Trihydroxybenzoic acid (3) | 100 | < 50.0 | - | - |
p-Hydroxycinnamic acid (4) | 100 | < 50.0 | - | - |
Ferulic acid (5) | 100 | 96.09 | 185.41 | |
50 | 64.53 | 35.97 | ||
10 | 28.33 | |||
Hirsutrin (6) | 100 | < 50.0 | - | - |
3′-Methoxyhirsutrin (7) | 50 | 85.89 | 64.04 | |
25 | 36.84 | 30.61 | ||
10 | 15.02 | |||
Cyanidin-3-glucoside (8) | 100 | < 50.0 | - | - |
Pelargonidin-3-glucoside (9) | 100 | 55.05 | 210.81 | |
50 | 13.35 | 91.36 | ||
10 | 1.09 | |||
Peonidin-3-glucoside (10) | 100 | < 50.0 | - | - |
Cyanidin-3-(6”-malonylglucoside) (11) | 50 | 91.73 | 54.06 | |
25 | 36.24 | 28.95 | ||
10 | 18.54 | |||
Peonidin-3-(6”-malonylglucoside) (12) | 100 | < 50.0 | - | - |
Kinetics of PTP1β inhibition by the active compounds
Optimization of extraction conditions to maximize total content of polyphenol, anthocyanin content, and extraction yield
No. | Independent | Response variables | |||||
---|---|---|---|---|---|---|---|
X1a | X2 | X3 | X4 | Y1b | Y2 | Y3 | |
1 | 30 (-1) | 6 (-1) | 25 (-1) | 12 (-1) | 5.91 | 55.16 | 6.00 |
2 | 50 (1) | 6 (-1) | 25 (-1) | 12 (-1) | 3.40 | 32.06 | 7.65 |
3 | 30 (-1) | 10 (1) | 25 (-1) | 12 (-1) | 6.55 | 65.20 | 6.00 |
4 | 50 (1) | 10 (1) | 25 (-1) | 12 (-1) | 5.47 | 47.47 | 6.70 |
5 | 30 (-1) | 6 (-1) | 35 (1) | 12 (-1) | 6.32 | 66.17 | 5.85 |
6 | 50 (1) | 6 (-1) | 35 (1) | 12 (-1) | 2.04 | 16.61 | 8.50 |
7 | 30 (-1) | 10 (1) | 35 (1) | 12 (-1) | 8.08 | 77.94 | 6.15 |
8 | 50 (1) | 10 (1) | 35 (1) | 12 (-1) | 3.70 | 34.71 | 7.30 |
9 | 30 (-1) | 6 (-1) | 25 (-1) | 12 (-1) | 4.42 | 41.02 | 6.30 |
10 | 50 (1) | 6 (-1) | 25 (-1) | 20 (1) | 6.64 | 67.21 | 7.75 |
11 | 30 (-1) | 10 (1) | 25 (-1) | 20 (1) | 6.33 | 63.36 | 6.25 |
12 | 50 (1) | 10 (1) | 25 (-1) | 20 (1) | 6.47 | 65.81 | 7.10 |
13 | 30 (-1) | 6 (-1) | 35 (1) | 20 (1) | 5.75 | 58.94 | 6.35 |
14 | 50 (1) | 6 (-1) | 35 (1) | 20 (1) | 3.62 | 70.81 | 9.25 |
15 | 30 (-1) | 10 (1) | 35 (1) | 20 (1) | 6.30 | 64.96 | 6.35 |
16 | 50 (1) | 10 (1) | 35 (1) | 20 (1) | 4.67 | 69.92 | 9.00 |
17 | 20 (-2) | 8 (0) | 30 (0) | 16 (0) | 3.41 | 20.63 | 5.75 |
18 | 60 (2) | 8 (0) | 30 (0) | 16 (0) | 3.80 | 24.07 | 9.10 |
19 | 40 (0) | 4 (-2) | 30 (0) | 16 (0) | 4.64 | 40.02 | 5.95 |
20 | 40 (0) | 12 (2) | 30 (0) | 16 (0) | 4.63 | 37.48 | 6.00 |
21 | 40 (0) | 8 (0) | 20 (-2) | 16 (0) | 6.22 | 52.90 | 6.40 |
22 | 40 (0) | 8 (0) | 40 (2) | 16 (0) | 4.92 | 71.46 | 5.75 |
23 | 40 (0) | 8 (0) | 30 (0) | 8 (-2) | 6.63 | 60.04 | 2.70 |
24 | 40 (0) | 8 (0) | 30 (0) | 24 (+2) | 5.72 | 56.32 | 6.35 |
25 | 40 (0) | 8 (0) | 30 (0) | 16 (0) | 4.95 | 45.34 | 5.75 |
26 | 40 (0) | 8 (0) | 30 (0) | 16 (0) | 4.34 | 35.13 | 5.78 |
27 | 40 (0) | 8 (0) | 30 (0) | 16 (0) | 4.28 | 40.88 | 5.73 |
Response variables | Second order polynomial equationsa | R2 | p-value |
---|---|---|---|
Total polyphenol content (mg GAE/g) | YTPC = 12.967917 + 0.236917X1 + 0.119583X2-0.184667X3-1.221042X4-0.001699X12-0.000781X1X2 + 0.021901X22-0.013988X1X3 + 0.003563 X2X3 + 0.012854X32 + 0.16953X1X42-0.021797X2X4-0.007281X3X4+ 0.029538X42 | 0.935 | 0.0001 |
Total anthocyanin content (mg C3GE/100 g) | YTAC = 412.389167 + 0.248000X1 + 4.349167X2-14.648667X3-23.737500X4-0.027356X12-0.058562X1X2 + 0.341094X22-0.079975X1X3-0.072875X2X3 + 0.288875X32 + 0.279406X1X42-0.230156X2X4 + 0.099687X3X4 + 0.388867X42 | 0.917 | 0.0001 |
Extraction yield (%) | YEY = 29.279167-0.538500X1-0.842500X2-0.823000X3-0.010833X4 + 0.006002X12-0.007188X1X2 + 0.059427X22 + 0.004625X1X3-0.003125X2X3 + 0.010508X32 + 0.004219X1X42 + 0.014844X2X4 + 0.003437X3X4-0.0077997X42 | 0.887 | 0.0001 |
Optimization of total polyphenol contents (TPC)
Response variables | Optimum extraction conditionsa | Maximum | Morphology | |||
---|---|---|---|---|---|---|
X1 | X2 | X3 | X4 | |||
Total polyphenol content (mg GAE/g) | 29.02 | 8.99 | 33.67 | 10.25 | 7.89 | Saddle point |
Total anthocyanin content (mg C3GE/100 g) | 45.22 | 7.85 | 34.14 | 22.97 | 82.44 | Saddle point |
Extraction yield (%) | 58.87 | 7.53 | 32.31 | 17.65 | 10.33 | Saddle point |
Response variables | F-radioa | |||
---|---|---|---|---|
X1 | X2 | X3 | X4 | |
Total polyphenol content (mg GAE/g) | 3.76 | 2.60 | 3.12 | 3.15 |
Total anthocyanin content (mg C3GE/100 g) | 1.90 | 4.29 | 2.83 | 2.98 |
Extraction yield (%) | 8.09 | 3.34 | 0.40 | 3.44 |
Optimization of total anthocyanin contents (TAC)
Response variables | Predicted values | Experimental values |
---|---|---|
Total polyphenol content (mg GAE/g) | 4.58 | 4.52 |
Total anthocyanin content (mg C3GE/100 g) | 42.28 | 43.02 |
Extraction yield (%) | 5.76 | 5.87 |