HPTLC Analysis of Bioactivity Guided Anticancer Enriched Fraction of Hydroalcoholic Extract of Picrorhiza kurroa

Table 4

(a) Robustness of the HPTLC method for estimation of cucurbitacins B, D, and E; betulinic acid; picrosides 1 and 2; and apocynin by changing detecting of wavelengths. (b) Robustness of the HPTLC method for estimation of cucurbitacins B, D, and E and betulinic acid by changing detecting of mobile phase composition. (c) Robustness of the HPTLC method for estimation of picrosides 1 and 2 and apocynin by changing detecting of mobile phase composition.

(a)

Parameters				Mean area ± SD^b	% RSD^c of area
	Components	Conc.; (ng spot^−1)	Wavelength used
Detecting wavelengths (nm^a) 240 and 595	Cucurbitacin B	100	238	554.33 ± 11.67	2.10
			242	545.66 ± 10.78	1.97
		200	238	875.66 ± 19.55	2.23
			242	876.66 ± 18.08	2.06
		400	238	1594.66 ± 25.71	1.61
			242	1588.66 ± 32.00	2.01
	Cucurbitacin D	200	238	441.0 ± 9.00	2.04
			242	438.33 ± 9.07	2.07
		400	238	830.33 ± 16.55	1.98
			242	824.33 ± 16.25	1.97
		800	238	1648.00 ± 32.42	1.97
			242	1650.33 ± 32.00	1.94
	Cucurbitacin E	200	238	640.66 ± 13.86	2.16
			242	638.00 ± 13.00	2.03
		400	238	1173.33 ± 22.03	1.94
			242	1169.33 ± 22.05	1.88
		800	238	2161.33 ± 41.24	1.91
			242	2154.33 ± 43.85	2.01
	Betulinic acid	250	593	981.33 ± 19.55	1.99
			597	977.33 ± 18.87	1.93
		500	593	1039.66 ± 21.00	2.02
			597	1031.00 ± 18.35	1.78
		1000	593	1363.66 ± 27.09	1.98
			597	1368.33 ± 27.61	2.01
Detecting wavelength (nm^a) 500	Picroside 1	100	498	941.33 ± 17.50	1.85
			502	946.00 ± 17.34	1.83
		200	498	1645.00 ± 34.05	2.06
			502	1644.33 ± 32.86	1.99
		400	498	2927.00 ± 55.05	1.88
			502	2940.00 ± 56.66	1.92
	Picroside 2	100	498	504.00 ± 10.00	1.98
			502	502.00 ± 9.53	1.90
		200	498	1004.00 ± 20.42	2.03
			502	1007.33 ± 19.50	1.93
		400	498	1919.33 ± 37.01	1.92
			502	1922.66 ± 40.69	2.11
	Apocynin	400	498	799.00 ± 15.50	1.94
			502	797.00 ± 14.54	1.82
		800	498	1593.33 ± 29.36	1.84
			502	1594.33 ± 31.65	1.98
		1000	498	2227.33 ± 40.50	1.81
			502	2257.00 ± 43.13	1.92

Nanometer. Standard deviation. Relative standard deviation.

(b)

Parameters			Mean area ± SD^a	% RSD^b of area
Components	Conc.; (ng spot^−1)	Mobile phase composition (toluene : ethyl acetate : formic acid, 60 : 40 : 0.5, v/v/v)
Cucurbitacin B	100	63 : 37 : 0.5	545.0 ± 11.78	2.16
		61 : 39 : 0.5	552.0 ± 12.00	2.17
		57 : 43 : 0.5	544.0 ± 10.44	1.91
	200	63 : 37 : 0.5	872.33 ± 16.28	1.86
		61 : 39 : 0.5	868.33 ± 16.25	1.87
		57 : 43 : 0.5	873.33 ± 16.56	1.91
	400	63 : 37 : 0.5	1605.0 ± 28.16	1.75
		61 : 39 : 0.5	1590.33 ± 26.33	1.67
		57 : 43 : 0.5	1603.66 ± 30.66	1.91
Cucurbitacin D	200	63 : 37 : 0.5	437.0 ± 9.00	2.06
		61 : 39 : 0.5	440.67 ± 9.5	2.16
		57 : 43 : 0.5	442.33 ± 10.07	2.28
	400	63 : 37 : 0.5	826.33 ± 15.04	1.82
		61 : 39 : 0.5	828.33 ± 17.01	2.05
		57 : 43 : 0.5	831.33 ± 15.72	1.89
	800	63 : 37 : 0.5	1644.00 ± 36.05	2.28
		61 : 39 : 0.5	1647.00 ± 29.82	1.81
		57 : 43 : 0.5	1643.66 ± 38.22	2.33
Cucurbitacin E	200	63 : 37 : 0.5	641.33 ± 12.34	1.92
		61 : 39 : 0.5	645.33 ± 12.01	1.86
		57 : 43 : 0.5	638.00 ± 12.53	1.96
	400	63 : 37 : 0.5	1176.67 ± 23.71	2.02
		61 : 39 : 0.5	1172.33 ± 22.59	1.93
		57 : 43 : 0.5	1170.33 ± 23.24	1.98
	800	63 : 37 : 0.5	2149.67 ± 40.55	1.89
		61 : 39 : 0.5	2154.0 ± 43.51	2.02
		57 : 43 : 0.5	2156.33 ± 44.0	2.04
Betulinic acid	250	63 : 37 : 0.5	978.0 ± 17.78	1.81
		61 : 39 : 0.5	977.33 ± 18.87	1.93
		57 : 43 : 0.5	972.33 ± 19.85	2.04
	500	63 : 37 : 0.5	1033.00 ± 21.79	2.10
		61 : 39 : 0.5	1022.66 ± 17.50	1.71
		57 : 43 : 0.5	1037.33 ± 18.92	1.82
	1000	63 : 37 : 0.5	1364.66 ± 25.69	1.88
		61 : 39 : 0.5	1371.00 ± 26.05	1.90
		57 : 43 : 0.5	1363.66 ± 27.09	1.98

Standard deviation. ^bRelative standard deviation.

(c)

Parameters			Mean area ± SD^a	% RSD^b of area
Components	Conc.; (ng spot^−1)	Mobile phase composition (toluene : ethyl acetate : methanol : formic acid, 40 : 50 : 10 : 0.5, v/v/v/v)
Picroside 1	100	42 : 48 : 10 : 0.2	946.00 ± 17.34	1.83
		38 : 52 : 10 : 0.2	948.33 ± 17.50	1.84
		40 : 52 : 08 : 0.2	948.66 ± 18.00	1.89
	200	42 : 48 : 10 : 0.2	1645.00 ± 34.04	2.06
		38 : 52 : 10 : 0.2	1643.00 ± 31.43	1.91
		40 : 52 : 08 : 0.2	1643.66 ± 30.66	1.86
	400	42 : 48 : 10 : 0.2	2927.00 ± 55.05	1.88
		38 : 52 : 10 : 0.2	2931.66 ± 60.87	2.07
		40 : 52 : 08 : 0.2	2927.66 ± 60.17	2.05
Picroside 2	100	42 : 48 : 10 : 0.2	504.00 ± 10.00	1.98
		38 : 52 : 10 : 0.2	501.33 ± 9.81	1.95
		40 : 52 : 08 : 0.2	506.66 ± 9.71	1.91
	200	42 : 48 : 10 : 0.2	1007.33 ± 19.50	1.93
		38 : 52 : 10 : 0.2	1005.66 ± 19.85	1.97
		40 : 52 : 08 : 0.2	1004.00 ± 20.42	2.03
	400	42 : 48 : 10 : 0.2	1914.33 ± 33.50	1.75
		38 : 52 : 10 : 0.2	1922.66 ± 40.68	2.11
		40 : 52 : 08 : 0.2	1915.33 ± 37.68	1.96
Apocynin	400	42 : 48 : 10 : 0.2	800.66 ± 15.94	1.99
		38 : 52 : 10 : 0.2	801.66 ± 15.63	1.94
		40 : 52 : 08 : 0.2	797.00 ± 14.52	1.82
	800	42 : 48 : 10 : 0.2	1593.33 ± 29.36	1.84
		38 : 52 : 10 : 0.2	1587.66 ± 28.67	1.80
		40 : 52 : 08 : 0.2	1597.00 ± 30.61	1.91
	1000	42 : 48 : 10 : 0.2	2222.66 ± 42.44	1.90
		38 : 52 : 10 : 0.2	2257.00 ± 43.13	1.91
		40 : 52 : 08 : 0.2	2228.00 ± 44.22	1.98

Standard deviation. Relative standard deviation.