Biotransformation of lignan glycoside to its aglycone by Woodfordia fruticosa flowers: quantification of compounds using a validated HPTLC method

Figures & data

Figure 1. Structures of lyoniside (1) and lyoniresinol (2).

Figure 2. UV spectra for lyoniside and lyoniresinol.

Figure 3. (a) Mass spectra for lyoniside. (b) Mass spectra for lyoniresinol.

Figure 4. (a) NMR spectra for lyoniside. (b) NMR spectra for lyoniresinol.

Table 1. Validation parameters for lyoniside and lyoniresinol.

Validation parameter	Lyoniside	Lyoniresinol
Linearity range (ng spot^−1)	250–3000	500–2500
Correlation coefficient (r^2)	0.9996; 0.9997	0.9988; 0.9994
Regression equation	6.9228 × −233.26	6.938 × −264.004
Limit of detection (ng spot^−1)	51.6	54.5
Limit of quantification (ng spot^−1)	152.2	165.8

Table 2. Intra- and inter-day precision for lyoniside and lyoniresinol.

	Lyoniside				Lyoniresinol
	Inter-day precision		Intra-day precision		Inter-day precision		Intra-day precision
Concentration (ng spot^−1)	Mean peak area±	% RSD	Mean peak area±	% RSD	Mean peak area±	% RSD	Mean peak area±	% RSD
500	3210.9 ± 34.11	1.06	3250.7 ± 46.99	1.44	3238.1 ± 35.27	1.089	3299.6 ± 56.68	1.71
1000	6204.23 ± 36.36	0.58	6412.15 ± 62.8	0.97	6442.19 ± 84.29	1.3	6385.4 ± 69.82	1.08
1500	10254.7 ± 90.92	0.88	10214.7 ± 63.73	0.62	10210.37 ± 81.99	0.8	10151.6 ± 117.6	1.15

Table 3. Robustness studies for lyoniside and lyoniresinol.

Change in mobile phase			Mean area ± SD		% RSD
Actual	Used	Level	LS	LR	LS	LR
4:3:0.4	3.5:3.5:0.4	−5	5995.9 ± 71.71	6339.46 ± 81.07	1.19	1.27
	4:3:0.4	0	6120.7 ± 62.07	6425.76 ± 60.96	1.01	0.94
	4.5:2.5:0.4	+5	6275.8 ± 78.45	6261.76 ± 72.53	1.25	1.15
Wavelength change
254	250	−4	6439.5 ± 60.71	6337.03 ± 68.46	0.94	1.08
	254	0	6357.8 ± 61.96	6432.36 ± 65.55	0.97	1.01
	258	+4	6441.8 ± 67.11	6268.03 ± 66.15	1.04	1.05

Table 4. Accuracy as recovery studies for lyoniside and lyoniresinol.

% of reference standard used for spiking	Theoretical content	Amount of drug recovered	% of drug recovered	% RSD
Lyoniside
0	2.5	2.49 ± 0.015	99.6	0.61
50	3.75	3.74 ± 0.02	99.73	0.53
100	5	5.01 ± 0.01	100.2	0.3
150	6.25	6.24 ± 0.03	99.84	0.48
Lyoniresinol
0	2.5	2.501 ± 0.011	100.04	0.46
50	3.75	3.73 ± 0.015	99.46	0.4
100	5	4.99 ± 0.03	99.8	0.64
150	6.25	6.23 ± 0.02	99.68	0.23

Figure 5. HPTLC chromatograms for aqueous extract and biotransformed extract (2–8 days).

Table 5. Quantification of lyoniside and lyoniresinol in aqueous extract and biotransformed product.

No. of days	LS	LR
0	13.98 ± 0.4	3.42 ± 0.06
1	13.54 ± 0.2	3.4 ± 0.04
2	13.44 ± 0.3	3.25 ± 0.03
3	12.2 ± 0.5	4.6 ± 0.02
4	10.3 ± 0.1	6.1 ± 0.05
5	6.4 ± 0.2	9.2 ± 0.02
6	5.3 ± 0.5	11.1 ± 0.04
7	4.3 ± 0.2	12.4 ± 0.02
8	3.7 ± 0.4	13.2 ± 0.03