Quantitative determination, principal component analysis and discriminant analysis of eight marker compounds in crude and sweated Dipsaci Radix by HPLC-DAD

Figures & data

Table 1. The origin and respective voucher number of Dipsaci Radix.

Samples	Origin	Voucher number
Crude drug	Sichuan-1	C20131209
	Sichuan-2	C20131209-1
	Sichuan-3	C20131209-2
	Sichuan-4	C20131209-3
	Sichuan-5	C20131006-3
	Hubei-1	C20131025
	Hubei-2	C20131006-4
	Hubei-3	C20131006-2
	Guizhou	C20131105-6
	Yunnan	C20131031
	Jiangxi	C20131006-5
Sweated sample	Sichuan-1	S20131209
	Sichuan-2	S20131209-1
	Sichuan-3	S20131209-2
	Sichuan-4	S20131209-3
	Sichuan-5	S20131006-3
	Hubei-1	S20131025
	Hubei-2	S20131006-4
	Hubei-3	S20131006-2
	Guizhou	S20131105-6
	Yunnan	S20131031
	Jiangxi	S20131006-5

Table 2. Linear relation between peak area and concentration.

Analytes	Regression equation	Correlation Coefficient (R^2)	Linear range (μg/mL)	LOD (μg/mL)	LOQ (μg/mL)
LA	y = 4.3712x − 39.286	0.9996	60.40–1208.00	0.15	0.50
CA	y = 1.9659x + 20.113	0.9998	151.00–3020.00	0.15	0.50
CaA	y = 31.195x + 40.929	0.9991	3.06–61.20	0.12	0.31
LN	y = 4.5192x − 1.2019	0.9998	30.76–615.20	0.12	0.31
I B	y = 32.669x + 0.4779	0.9998	5.13–102.60	0.15	0.51
IA	y = 16.765x + 30.678	0.9996	10.17–203.40	0.10	0.31
IC	y = 23.963x − 7.5264	0.9998	10.20–204.00	0.10	0.31
AVI	y = 1.7764x − 3.7019	0.9998	151.60–3032.00	0.15	0.50

Table 3. Precision of analytical results (n = 6).

		Intra-day	Inter-day
Analytes	Standard solution (μg/mL)	Precision (RSD/%)	Precision (RSD/%)
LA	120.80	0.71	1.24
	241.60	0.67	1.21
	604.00	0.62	1.18
CA	302.00	0.86	1.02
	604.00	0.52	1.43
	1510.00	0.50	0.94
CaA	6.12	1.10	1.77
	12.24	0.83	1.66
	30.60	0.52	1.32
LN	61.52	1.04	1.58
	123.04	0.85	1.53
	307.60	0.80	1.39
IB	10.26	0.86	1.22
	20.52	0.86	1.35
	51.30	0.51	1.18
IA	20.34	0.79	0.98
	40.68	0.62	1.58
	101.70	0.59	1.54
IC	20.40	0.61	1.52
	40.80	0.76	1.31
	102.00	0.77	0.98
AVI	303.20	0.53	1.17
	606.40	0.83	1.27
	1516.00	0.77	0.96

Table 4. Repeatability and stability of analytical results (n = 6).

	Repeatability (% ± SD, RSD/%)		Stability (% ± SD, RSD/%)
Analytes	Crude drug	Sweated sample	Crude drug	Sweated sample
LA	0.86 ± 0.02, 2.02	0.71 ± 0.02, 2.73	0.87 ± 0.01, 1.21	0.72 ± 0.01, 1.29
CA	2.54 ± 0.04, 1.75	2.33 ± 0.05, 2.28	2.51 ± 0.04, 1.56	2.39 ± 0.04, 1.61
CaA	0.11 ± 0.00, 2.23	0.11 ± 0.00, 3.44	0.10 ± 0.00, 1.57	0.11 ± 0.00, 1.97
LN	0.57 ± 0.01, 1.90	0.53 ± 0.02, 3.45	0.57 ± 0.01, 2.15	0.54 ± 0.01, 1.82
IB	0.12 ± 0.00, 1.64	0.13 ± 0.00, 1.95	0.12 ± 0.00, 1.42	0.13 ± 0.00, 1.86
IA	0.59 ± 0.02, 2.60	0.58 ± 0.01, 2.55	0.59 ± 0.01, 1.05	0.58 ± 0.01, 1.43
IC	0.38 ± 0.01, 2.53	0.48 ± 0.02, 3.44	0.37 ± 0.01, 1.42	0.48 ± 0.00, 0.85
AVI	6.26 ± 0.15, 2.35	5.37 ± 0.08, 1.40	6.25 ± 0.06, 0.89	5.39 ± 0.07, 1.22

Table 5. Recoveries of marker compounds through standard addition (n = 6).

Analytes	Original (mg ± SD)	Spiked (mg)	Determined (mg ± SD)	Recovery (% ± SD, RSD%)
LA	2.03 ± 0.00	1.60	3.66 ± 0.07	101.86 ± 4.12, 4.05
		2.00	4.05 ± 0.05	100.91 ± 2.75, 2.73
		2.40	4.46 ± 0.07	101.24 ± 2.84, 2.80
CA	5.93 ± 0.01	4.80	10.75 ± 0.14	100.39 ± 2.84, 2.83
		6.00	12.01 ± 0.23	101.43 ± 3.78, 3.73
		7.20	13.07 ± 0.21	99.22 ± 3.01, 3.03
CaA	0.25 ± 0.00	0.20	0.45 ± 0.01	100.47 ± 3.84, 3.82
		0.25	0.50 ± 0.01	100.38 ± 3.96, 3.94
		0.30	0.55 ± 0.01	99.76 ± 3.63, 3.64
LN	1.23 ± 0.00	0.96	2.21 ± 0.02	102.35 ± 1.94, 1.89
		1.20	2.42 ± 0.03	99.38 ± 2.57, 2.59
		1.44	2.70 ± 0.05	102.26 ± 3.33, 3.26
IB	0.27 ± 0.00	0.20	0.47 ± 0.01	99.91 ± 2.70, 2.70
		0.25	0.53 ± 0.01	101.26 ± 2.05, 2.02
		0.30	0.57 ± 0.01	99.94 ± 3.95, 3.95
IA	1.34 ± 0.00	1.08	2.45 ± 0.03	102.18 ± 2.63, 2.57
		1.35	2.68 ± 0.04	99.03 ± 2.68, 2.71
		1.62	3.00 ± 0.04	102.38 ± 2.17, 2.12
IC	0.87 ± 0.00	0.70	1.57 ± 0.02	99.84 ± 3.05, 3.04
		0.85	1.71 ± 0.02	99.28 ± 2.19, 2.21
		1.02	1.91 ± 0.03	102.01 ± 3.17, 3.10
AVI	14.43 ± 0.02	11.60	26.23 ± 0.24	101.72 ± 2.17, 2.14
		14.50	29.06 ± 0.48	101.29 ± 2.81, 2.77
		17.40	32.21 ± 0.54	102.21 ± 3.16, 3.09

Figure 1. HPLC chromatogram of standard mixture (A), crude drug (B) and sweated sample (C): (1) LA; (2) CA; (3) CaA; (4) LN; (5) IB; (6) IA; (7) IC and (8) AVI.

Figure 2. ESI-MS spectra of eight compounds.

Table 6. Contents of eight components in Dipsaci Radix (% ± SD, n = 3).

Suppliers	Samples	LA	CA	CaA	LN	IB	IA	IC	AVI
Sichuan-1	Crude drug	1.10 ± 0.03	8.28 ± 0.21	0.01 ± 0.00	0.61 ± 0.03	0.09 ± 0.00	0.92 ± 0.03	0.48 ± 0.02	15.19 ± 0.34
	Sweated sample	0.98 ± 0.04	2.49 ± 0.11	0.05 ± 0.00	0.45 ± 0.02	0.14 ± 0.01	0.70 ± 0.03	0.59 ± 0.02	2.50 ± 0.12
	change rate	−10.91%	−69.93%	+400.00%	−26.23%	+55.56%	−23.91%	+22.92%	−83.54%
Sichuan-2	Crude drug	1.10 ± 0.04	8.28 ± 0.22	0.01 ± 0.00	0.61 ± 0.03	0.09 ± 0.00	0.92 ± 0.04	0.48 ± 0.02	15.19 ± 0.35
	Sweated sample	0.94 ± 0.03	4.11 ± 0.21	0.09 ± 0.00	0.51 ± 0.02	0.18 ± 0.01	0.68 ± 0.03	0.80 ± 0.02	2.31 ± 0.11
	change rate	−14.55%	−50.36%	+800.00%	−16.39%	+100.00%	−26.09%	+66.67%	−84.79%
Sichuan-3	Crude drug	1.10 ± 0.03	8.28 ± 0.25	0.01 ± 0.00	0.61 ± 0.02	0.09 ± 0.00	0.92 ± 0.03	0.48 ± 0.01	15.19 ± 0.42
	Sweated sample	0.93 ± 0.02	2.99 ± 0.12	0.07 ± 0.00	0.43 ± 0.02	0.16 ± 0.01	0.55 ± 0.02	0.55 ± 0.02	1.95 ± 0.09
	change rate	15.45%	−63.89%	+600.00%	−29.51%	+77.78%	−40.22%	+14.58%	−87.16%
Sichuan-4	Crude drug	1.10 ± 0.04	8.28 ± 0.26	0.01 ± 0.00	0.61 ± 0.02	0.09 ± 0.00	0.92 ± 0.04	0.48 ± 0.02	15.19 ± 0.38
	Sweated sample	0.96 ± 0.03	2.61 ± 0.13	0.13 ± 0.01	0.49 ± 0.02	0.12 ± 0.01	0.59 ± 0.02	0.55 ± 0.02	1.94 ± 0.06
	change rate	−12.73%	−68.48%	+1200.00%	−29.51%	+77.78%	−35.87%	+14.58%	−87.23%
Sichuan-5	Crude drug	0.89 ± 0.04	7.62 ± 0.08	0.01 ± 0.01	0.54 ± 0.02	0.08 ± 0.01	0.79 ± 0.02	0.38 ± 0.01	6.33 ± 0.21
	Sweated sample	0.68 ± 0.03	2.29 ± 0.06	0.11 ± 0.01	0.42 ± 0.01	0.13 ± 0.01	0.58 ± 0.02	0.46 ± 0.02	5.31 ± 0.17
	change rate	−23.60%	−69.95%	+1000.00%	−22.22%	+62.50%	−26.58%	+14.58%	−16.11%
Hubei-1	Crude drug	0.86 ± 0.04	2.57 ± 0.08	0.16 ± 0.00	0.57 ± 0.02	0.11 ± 0.01	0.67 ± 0.03	0.64 ± 0.01	1.61 ± 0.06
	Sweated sample	0.68 ± 0.03	2.23 ± 0.07	0.18 ± 0.01	0.54 ± 0.01	0.13 ± 0.01	0.61 ± 0.03	0.78 ± 0.03	1.36 ± 0.04
	change rate	−20.93%	−13.23%	+12.50%	−5.26%	+18.18%	−8.96%	+17.95%	−15.53%
Hubei-2	Crude drug	0.83 ± 0.03	6.85 ± 0.24	0.02 ± 0.00	0.51 ± 0.01	0.08 ± 0.00	0.73 ± 0.03	0.63 ± 0.03	4.99 ± 0.21
	Sweated sample	0.54 ± 0.02	5.00 ± 0.15	0.03 ± 0.00	0.39 ± 0.01	0.09 ± 0.00	0.60 ± 0.02	0.73 ± 0.03	3.58 ± 0.13
	change rate	−34.94%	−27.01%	+50.00%	−23.53%	+12.50%	−17.81%	+15.87%	−28.26%
Hubei-3	Crude drug	1.01 ± 0.04	4.38 ± 0.09	0.10 ± 0.00	0.48 ± 0.01	0.12 ± 0.01	0.88 ± 0.03	0.48 ± 0.02	9.59 ± 0.27
	Sweated sample	0.81 ± 0.03	4.25 ± 0.18	0.11 ± 0.01	0.47 ± 0.01	0.13 ± 0.01	0.81 ± 0.03	0.58 ± 0.02	9.31 ± 0.32
	change rate	−19.80%	−2.97%	+10.00%	−2.08%	+8.33%	−7.95%	+20.83%	−2.92%
Jiangxi	Crude drug	0.81 ± 0.04	6.79 ± 0.21	0.03 ± 0.00	0.49 ± 0.02	0.08 ± 0.00	0.74 ± 0.02	0.73 ± 0.03	4.78 ± 0.15
	Sweated sample	0.53 ± 0.02	4.54 ± 0.17	0.04 ± 0.00	0.38 ± 0.01	0.10 ± 0.00	0.63 ± 0.02	0.78 ± 0.03	3.43 ± 0.12
	change rate	−34.57%	−33.14%	+33.33%	−22.45%	+25.00%	−14.86%	+14.58%	−28.24%
Guizhou	Crude drug	0.71 ± 0.03	2.02 ± 0.07	0.03 ± 0.00	0.29 ± 0.01	0.08 ± 0.00	0.77 ± 0.02	0.26 ± 0.01	4.74 ± 0.22
	Sweated sample	0.50 ± 0.02	1.67 ± 0.06	0.04 ± 0.00	0.21 ± 0.01	0.09 ± 0.00	0.75 ± 0.03	0.27 ± 0.01	4.16 ± 0.15
	change rate	−29.58%	−17.33%	+33.33%	−27.59%	+12.50%	−2.60%	+3.85%	−12.24%
Yunnan	Crude drug	0.79 ± 0.03	2.54 ± 0.08	0.06 ± 0.00	0.48 ± 0.02	0.06 ± 0.00	0.66 ± 0.02	0.35 ± 0.01	2.83 ± 0.12
	Sweated sample	0.66 ± 0.02	1.89 ± 0.07	0.07 ± 0.00	0.38 ± 0.01	0.07 ± 0.00	0.48 ± 0.02	0.44 ± 0.02	2.14 ± 0.08
	change rate	−16.46%	−25.59%	+16.67%	−20.83%	+16.67%	−27.27%	+25.71%	−24.38%

“−”decreased; “+” increased.

Figure 3. Principal component analysis (PCA) of Dipsaci Radix.

Figure 4. Discriminant analysis (DA) of the crude and sweated Dipsaci Radix.